M2M+Unit+III+LOs

toc

=Molecular Basis of Carcinogenesis I, II, III= [These are a little scattered] [Notice that the notes have the following unelaborated steps: Tumor initiation, promotion, conversion, progression.]
 * Describe the properties of malignant cancer cells. You should be able to give at least five different properties.
 * Phenotype properties:
 * Unresponsive to normal signals for proliferation control.
 * De-differentiated (lack specialized structures of tissues in which they grow).
 * Invasive (can grow out into neighboring tissues).
 * Metastatic (can shed into circulation and proliferate elsewhere in the body).
 * Generally it's the metastasis that's fatal in cancers.
 * Clonal origin (derived from a single cell).
 * Increased transport of glucose.
 * Lack of contact inhibition (will grow over each other).
 * Immortality (ability to grow indefinitely).
 * Can grow without an attachment to a solid substrate.
 * What is the multi-step process for carcinogenesis? You should be able to discuss the relative importance of heredity and the environment and why early events may include mutations in DNA repair genes.
 * Not considered to be an inherited disease (not inherited as a single Mendelian gene).
 * On the other hand, cancer __susceptibility__ genes are certainly heritable (see Knudson below).
 * Carcinogenesis is characterized by the accumulation of many genetic alterations or mutations, particularly over a long period of time-- thus age is strongly associated with cancer, as are environmental factors that produce high rates of mutations.
 * If DNA repair genes are damaged early on, the rate at which you accumulate DNA mutations - since you can't repair them as well - goes up markedly.
 * Multi-step process for cancer:
 * (1) Normal cell
 * (2) Increased proliferation: With a mutation or two, an immortalized cell (see below for examples).
 * (3 + 4) Early/progressive neoplasia: With a few more mutations, get abnormal growth patterns.
 * (5) Carcinoma: A full-on tumor.
 * (6) Metastasis: A tumor that's spreading through the circulatory system.
 * I think the point he's trying to make here is that you need a fairly wide assortment of mutations (though order isn't important) to result in a cancer:
 * Turn on oncogene or make oncogene protein much more active
 * Turn off tumor suppressor genes (both cell cycle regulatory and DNA repair genes)
 * Turn off apoptotic genes and turn on anti-apoptotic genes.
 * What types of genes are usually mutated in tumor initiation? Describe the effect on cellular proliferation that the product of these genes has.
 * Either activated oncogenes (proliferation genes) or silenced tumor suppressor genes.
 * Oncogenes: accelerate proliferation. Tumor suppressors: slow down proliferation.
 * What type of cytogenetic abnormalities are associated with malignancy? You should be able to give at least two different examples.
 * Translocations of chromosomes, deletions on chromosomes:
 * Can activate oncogenes (for example, by putting an extremely active promoter upstream of one).
 * Can inactivate tumor suppressor genes (for example, by translocating another gene into the middle of the tumor suppressor sequence)
 * Notice that this is kind of a silly point to make, at least on its face. Anything that can bring a promoter nearby an oncogene (like pretty much any chromosomal rearrangement) or anything that can interrupt transcription or promotion of a tumor suppressor gene (likewise) can be associated with malignancy.
 * Notice that aneuploidy (recall: loss or gain of a chromosome) is associated with a poor outcome in many cancers.
 * What events can produce LOH? Give at least two examples and state how they support Knudson’s theory.
 * LOH = Loss Of Heterozygosity.
 * Means you inherit a heterozygous state (say, for a working tumor suppressor gene), but convert to (negative) homozygosity at some point.
 * Loss of this heterozygosity means you lose the one working copy of the gene that you have, through:
 * mutation
 * mitotic recombination
 * chromosome loss
 * and/or environmental factors
 * Knudson sez: If you're heterozygous, you have one "strike" against you through your genes (one copy of suppressor gene knocked out in your parents' passed-on DNA). If you have one more "strike" (ie, exposure to UV light causes an unrepaired mutation in the other copy of that gene), you're unable to produce that tumor suppressor gene product at all-- which leads, potentially, to cancer.
 * Example: Familial retinoblastoma vs acquired retinoblastoma (in children vs. adults, bilateral vs. unilateral)-- former is easier to acquire since it only requires one mutation event.
 * Are cancers associated with both dominant and recessive syndromes? You should be able to give a different example of each type.
 * Familial retinoblastoma: A recessive disease but inherited in a dominant way: need both tumor suppressor (RB) genes knocked out to show a phenotype (thus recessive), but everyone who inherits heterozygosity winds up with the disease (thus dominant) due to LOH problems.
 * Notice that this inheritance pattern shows a vertical pedigree (looks like autosomal dominant).
 * Sporadic retinoblastoma: Mutations in both tumor suppressor genes needed to knock out function- thus not strongly inherited (not sure how he's tying this to the LO, as somatic sporadic mutations shouldn't be inherited at all).
 * Describe how the RB (retinoblastoma) gene was first identified. You should be able to describe the important cytogenetic and molecular evidence.
 * Sorry, I was scrambling around trying to figure out the last thing he said when he was going over this. It probably has something to do with examining pedigrees of familial RB patients, noticing the inheritance pattern, and using some kind of tagging technology to figure out which gene was present in those with a greater tendency to get RB and absent in those without that tendency.
 * What are the properties of the protein product of the RB gene? List at least three biochemical properties
 * [Notice that the RB protein is a universal protein-- not just found in the retina.]
 * An inhibitor of the cell cycle that prevents proliferation.
 * Normally __hypophosphorylated__ (little PO4) to prevent proliferation.
 * __Hyperphosphorylated__ by __CDKs__ (cyclin-dependent kinases) to be turned __off__.
 * When turned __off__, allows normal cell proliferation.
 * When __kept__ off or inhibited, allows unchecked cell proliferation and carcinogenesis.
 * General note: RB protein works by binding to a variety of transcription factors.
 * Describe how the RB protein functions during the cell cycle and why it is important in cancer. You should be able to give an explanation of how the loss of RB may produce a malignancy.
 * As mentioned, functions to block G1 moving to S (replication) phase. Without RB, a cell has no 'brake' on its proliferation.
 * Notice that there are certain tissues that are particularly susceptible to the loss of RB-- that is, losing RB there has a particularly acute effect. Example is, obviously, the retina.
 * What is the hallmark of a tumor suppressor gene or anti-oncogene? You should be able to use the RB gene as an example
 * Prevents cells from proliferating by controlling cell cycle.
 * How were oncogenes discovered? You should be able to describe the method with at least three different examples
 * Discovered in oncogenic retroviruses (specifically Rous Sarcoma Virus in chickens). With one particular viral gene segment ( // v-onc // ), tumors are rapidly induced in the infected cells after infection; without it, integration into the host genome occurs without activation of oncogenes.
 * Examples: // v-src //, // v-erb // , // v-myc // , etc. Watch for the // v- // at the beginning of it. (as opposed to // c-myc // , which is an endogenous oncogene in the human genome.)
 * Often oncogenes mimic growth factor receptors to achieve their nefarious ends.
 * Method: take cells, put them in agar, watch for proliferation. Normal cells won't be able to proliferate (no anchorage to grow on)-- infected cells will proliferate regardless of anchorage (see characteristics of cancer cells, above).
 * What functions do the protein products of viral oncogenes perform? You should be able to give at least four examples of oncogenes of known function
 * RB protein is a target of some tumor viruses (eg. human papilloma virus), which produce proteins which inactivate RB (and/or p53) in the cell in which the virus has taken up residence.
 * This is a common theme: RB and/or p53 inactivation by viral proteins allowing rapid, unchecked cell proliferation.
 * Notice Karposi's sarcoma [HIV/AIDS] is also caused by RB/p53-inactivating proteins.
 * Notice also that __retro__viral-induced cancers (ie, resultant from viral reverse-transcription of their oncogenes into human DNA) are very rare in humans. Our viruses tend to just inactivate p53 and RB rather than encode oncogenes themselves.
 * Oncogenes:
 * As mentioned, the viral // src //, // erb // , // myc // , etc, sequences. Notice that viral copies of oncogenes tend to be more powerful effects than their endogenous counterparts.
 * // v-src // : phosphorylates various tyrosine residues in other proteins (similar to ABL in humans).
 * // v-erb // : mimics epidermal growth factor receptor (unregulated).
 * // v-sis // : mimics platelet-derived growth factor (unregulated).
 * Endogenous oncogenes are marked // c-onc // :
 * Notice that // c-onc // genes are part of normal functioning of human cells; therapy can't target all // c-onc //, just their overexpression.
 * // c-onc // genes need to undergo mutation before they become carcinogenic.
 * [Not on LOs but probably helpful:]
 * __APC__ gene product: keeps beta-catenin outside the nucleus; without APC, beta-catenin goes to nucleus and begins uncontrolled transcription of oncogenes (like // c-myc // ).
 * LOH in APC produces familial adenomatous polyposis (FAP), which leads to colon polyps and, eventually, metastatic colon cancer.
 * __BRCA1__ (breast-cancer gene 1): Its gene product forms the scaffold for protein assembly that "checks up on" the cell cycle to make sure that the DNA has replicated faithfully. When it's knocked out, the check on the cell cycle is removed.
 * Can either be familial (LOH as above) or sporadic.
 * Note that in sporadic cases, the mutation can be on other (unspecified) genes that regulate or have an effect on BRCA's expression.
 * __p53__ you may recall from Li-Fraumeni syndrome-- mutant or inactivated p53 is found in ~50% of all cancers.
 * Protein involved in DNA mutation repair at "checkpoint" in cell cycle.
 * Notice that mutations in p53 can be "dominant-negative;" that is, one bad copy of p53 can inactivate the other, good copy of p53.
 * Along with RB, one of the two major gene products knocked out by oncoviruses to produce cancers.
 * Why are oncogenes useful as molecular markers in prognosis? You should be able to give at least two examples of oncogenes that are currently being used and also include the evidence of why these are good markers.
 * The level of expression of oncogenes tends to correlate with the rapidity of the progress of the cancer.
 * One example: The level of expression of the // N-myc // gene is used in prognosis analysis for neuroblastoma.
 * Another example: Increased expression of the // HER2/neu // gene correlates with poor prognosis in breast cancer.
 * What is the difference between oncogenes and tumor suppressor genes? You should be able to describe the function of these two types of cancer genes and how mutations in them may combine to produce cancers.
 * Again: oncogenes promote cell proliferation, tumor suppressor genes inhibit it. If you have an over-activation mutation in an oncogene and an inhibiting mutation in a tumor suppressor gene, presumably you can have a real problem.
 * How can our knowledge of oncogenes and tumor suppressor genes be used for targeted therapy? You should be able to give an example of each.
 * Can design either small molecules or antibodies as therapy.
 * Targeting oncogenes:
 * Herceptin: drug antibody therapy against the HER2/erb2 oncogene product.
 * Small molecules: able to inhibit action of cancerous proteins by, among other pathways, binding to their active sites. Example is Gleevac, an ATP analogue, that inhibits ABL tyrosine kinase in patients with BRC-ABL translocation on the Philadelphia chromosome.
 * (BRC-ABL: translocation causing leukemia; resistant to radiation therapy. ABL is an ATP-dependent tyrosine protein kinase that PO4s various other proteins.)
 * Gleevac = specific to tyrosine kinase proteins: specific fit to their ATP-binding pocket.
 * Notice can use combined oncogene targeting therapy and radiation therapy: former makes the cancer more susceptible to the radiation.
 * Targeting tumor repressor genes:
 * You can inject RB directly into RB-negative tumors
 * You can use drugs that only kill cells with p53 deficiencies
 * You can use drugs which correct the mutant conformation of dominant-negative p53 proteins (see above).

=Composition of Cells=
 * Know typical values for the volumes of plasma (**3 liters** ), extracellular fluid ( **ECF, about 13 liters** + **another 5 for the 'third space'** ), and intracellular fluid ( **ICF, about 27 liters** ) compartments.
 * Of the roughly 45 liters of fluid in the body:
 * 99% is H2O
 * 0.8% is Na+, K+, Cl-
 * 0.2% is everything else.
 * Notice that there's roughly a 2:1 ratio between ICF and ECF.
 * 'Third space': fluid in GI tract, urine, CSF, sweat, etc. Effectively this is any fluid outside of cells but separated from interstitial fluid by epithelial cells. At the moment, more or less irrelevant.


 * Know the major difference in composition between ECF and ICF.
 * In ICF: mostly K+ cations (98% of K+ is inside cells) and a variety of anions (A-n, where -n is the average charge on the anions).
 * In ECF: mostly Na+ cations and Cl- anions.
 * [Types of ECF: interstitial fluid, lymph, plasma]
 * [Types of ICF: mitochondrial, nuclear, endoplasmic reticular, etc]
 * [For purposes of discussion, we're lumping all these together into ICF and ECF.]
 * Normal ion values:
 * __ICF__ (mM)
 * __ECF__ (mM)
 * __Membrane Permeability__ (yes/no)
 * Na+
 * 14
 * 140
 * (no) (yes, but actively pumped out)
 * K+
 * 145
 * 5
 * yes
 * Cl-
 * 5
 * 145
 * yes
 * A-n (anions)*
 * 126
 * ~0
 * no
 * H2O
 * ~55,000
 * ~55,000 (same as ICF)
 * yes
 * * = proteins, inorganic ions, etc.
 * Know the two most important functional properties of membranes, one conveyed by lipids, the other by channels and transporters.
 * Lipids are resistant to charge diffusion: that is, they can hold a differential charge on either side without having the charge leak across and balance out. There are two components to this:
 * One is that they are __impermeable to charged substances__ (ie water and ions).
 * The other is that they are 'strong' vs. electric force: they can withstand the large amount of electric force imposed by the charge differentiation. (note that if you increase the membrane differential by another 50 mV or so, this breaks down.)
 * Channels and transporters (proteins in lipid membrane) are the ways by which a cell gets around this impermeability. This allows a cell to be __selectively permeable__ and move ions back and forth in a controlled fashion.
 * // Channels // : allows passive diffusion out of or into cell. Notice:
 * They are generally __selective__ for a particular type of ion, and
 * They're usually '__gated__'-- that is, there's a particular type of stimulus that will cause it to open (particular receptor-binding ligands, stretch receptors, voltage changes, etc).
 * // Transporters // : actively pump ions out of or into cell.
 * Generally selectively bind large molecules for transport.
 * Can also pump ions against a gradient (ie. Na+ pump) using ATP.
 * Understand the routes by which a given substance can traverse a membrane.
 * See last two points (channels and transporters).
 * Notice, peripherally, that if the substance is lipid-soluble, it can just go straight into (and potentially through) the membrane itself (how it would have gotten to the membrane through the external aqueous environment is another question).
 * Identify physical forces that can determine the gating properties of ion channels.
 * Electric field (voltage-gated)
 * Mechanical (stretch, hair cells in cochlea)
 * Chemical (synaptic receptors)
 * Temperature (cutaneous temp receptors)

=Cell Volume Regulation=
 * Be able to determine which direction a given uncharged substance will move across a membrane, given its concentrations on the two sides.
 * Basic law of everything: substances tend to move from areas of high concentration to areas of low concentration. This probably is connected to the second law of thermodynamics (entropy, or randomness, or diffusion, is constantly increasing).
 * Notice that the diffusion of water down a gradient (ie. towards the area of low concentration) is called __osmosis__. This usually is discussed in the context of water flowing through a membrane that's impermeable to other substances, to balance out uneven osmotic concentrations of those concentrations on either side of the membrane.
 * Determine under a given set of conditions whether a cell will swell or shrink.
 * Notice that, given the relative concentrations of water vs. everything else inside of (and outside of) cells, __water__ alone is effectively going to determine changes in the volume of cells.
 * If the cell is immersed in water that's at a higher concentration (that is, it's more pure water and less solute) than the water inside the cell, the cell will __swell__.
 * If the cell is immersed in water that's at a lower concentration (that is, it's less pure water and more solute) than the water inside the cell, the cell will __shrink__.
 * Notice there's another way of saying this:
 * If the cell is immersed in water that has a __lower__ concentration of non-water stuff than the concentration of non-water stuff __inside__ the cell, the cell will __swell__ (water flows into cell to balance the concentrations).
 * If the cell is immersed in water that has a __higher__ concentration of non-water stuff than the concentration of non-water stuff __inside__ the cell, the cell will __shrink__ (water flows out of cell to balance the concentrations).
 * Notice that in pure water, our cells will burst (too much swelling!) because they're always going to have a higher concentration of solutes than their surroundings.
 * List the three mechanisms that different cells have evolved to keep from swelling and bursting.
 * (1) Make cell membranes impermeable to water.
 * This is not a popular option, since during cell division you're effectively going from 1 cell-full of water to 2 cell-fulls, and you've got to get water in there somehow to make up the extra volume. In other words: cells need to grow.
 * However, this can work sometimes in specialized systems (ie kidney cells when you need to retain water).
 * (2) "Brute force": build a wall around the cell which resists the osmotic pressure.
 * This is by far the most popular option-- all plant matter and pretty much everything other than animal cells.
 * The wall is permeable to water but resists the osmotic swelling force.
 * One problem is that it really limits mobility and morphology of the cell. The other problem is that it takes a lot of uptake and energy to maintain that wall.
 * (3) Balance the water concentrations osmotically.
 * Ie: put solute in extracellular fluid to make the water outside cells the same concentration as the water inside cells.
 * Notice that it doesn't really matter what solute you put in the ECF to dilute the water, as long as the solutes aren't able to go into the ICF (which would render the whole point moot).
 * Know which direction water will move across a semi-permeable membrane, given the solute compositions of the fluids on either side of the membrane.
 * Detailed answer: it depends on whether or not those solutes can cross the membrane and how fast they do so. See below, under "reflection coefficients," for details. Essentially if you have one solute that permeates the membrane more quickly than the other, the water will follow that solute first. If you have two permeable solutes, one on each side of the membrane, that both diffuse at the same rate, then the net water flow should be nada.
 * Simple answer: if the solutes can't cross the membrane, water will flow in the direction of greater solute concentration.
 * Know the difference between diffusion and osmosis.
 * Osmosis is the net inward movement of water across a semi-permeable membrane. Notice more detail in the definition on the first bullet point.
 * Diffusion is the more general movement of any solute from an area of high concentration of that solute to an area of low concentration of that solute.
 * Know which substances must move into or out of a cell in order for the cells volume to change.
 * Water alone is necessary and sufficient. To a first approximation.
 * Know the effect of having a membrane with different, non-zero permeabilities (ie, reflection coefficients less than 1) to different solutes.
 * ['reflection coefficient' = how permeable the cell membrane is to a substance: 0 = water, 1 = impermeable.]
 * Effectively, some things pass into cells more quickly than others, some things can't come into cells at all.
 * This has consequences: given the same concentrations of different __diffusable__ solutes on each side of the membrane, until they equilibrate out, one of them will likely diffuse __faster__ than the other one, which means that the balance of solute will __not__ be balanced throughout the diffusion process (which means the cell is going to either swell or shrink in the process as water follows the solute that diffuses faster).
 * Know the definitions of molarity, osmolarity, equivalents, and tonicity, and know how to convert between them.
 * Molarity: a standard measure of __concentration__ (grams of solute per liter of water).
 * Osmolarity: sum of molarity of all the solute particles. Effectively, you want to (ie. 1 M NaCl = 2 OsM since you have 1 M of Na+ and 1 M of Cl- when dissolved). Can calculate this for inside and outside the cell and compare them.
 * Notice that 1 M glucrose (C6H12O6) doesn't wind up being something crazy like 24 OsM, even though there's 24 atoms in each glucose molecule, how many atoms are in each, because sucrose doesn't dissolve into constituent atoms in water like ions in a salt do.
 * This can be used to determine whether a cell will swell or shrink, as noted above. Basically if you have a __higher__ osmolarity outside the cell than inside, the cell will __shrink__. If you have a __lower__ osmolarity outside the cell than inside, the cell will __swell__. If the osmolarity is the same inside and outside, the cell will stay the same size.
 * Important point: in a simplified situation (not comparing reflection coefficients or two opposing diffusing solutes), a solute that can cross the lipid membrane __does not change the volume of a cell at all__. It will go to equilibrium across that membrane, which means that the osmolarity both inside and outside the cell increases by the same amount, which means the __difference__ in osmolarity doesn't change at all-- thus no volume change.
 * But note caveat about relative permeabilities, above.
 * Tonicity: a measure of how a cell reacts to a given ECF solution. Also a way of describing the osmolarity of the __impermeable__ solutes in the ECF.
 * __Hypotonic__: any solution that makes a cell __swell__. (total __impermeable__ osmolarity = lower outside the cell)
 * __Hypertonic__: any solution that makes a cell __shrink__. (total __impermeable__ osmolarity = higher outside the cell).
 * __Isotonic__: any solution that doesn't make a cell shrink or swell- same __impermeable__ osmolarity as what's in the cell.
 * Equivalents: Take the ions that are the solute, break them down into osmolarity as above, then multiply the osmolarity by the charge on the ion. Basically gives you a feel for how much of a given osmolarity is acidic or basic. I find it unlikely he'll ask a question about this, but just in case.

=Membrane Potential I, II=
 * Compare qualitatively the relative strengths of electric and osmotic forces:
 * Electric forces are way crazy bigger than osmotic forces.
 * This is why a small number of ions can overcome a large concentration gradient.
 * Describe the two forces acting on an ion moving across a membrane:
 * Concentration gradient (higher concentrations diffuse to lower)
 * Electrical gradient (like charges repulse, opposite charges attract)
 * Define equilibrium potential:
 * For a given ion, the equilibrium potential is the potential across the membrane that that ion would 'like' to see exist in order to be in equilibrium.
 * If the membrane potential isn't at the equilibrium potential for the given ion, either the ion isn't able to permeate the membrane or there's some mechanism that keeps the ions from crossing the membrane effectively in order to equilibrate (ie, an ion pump).
 * Measured in millivolts.
 * Understand the difference between an equilibrium potential and a recorded membrane potential:
 * Membrane potential = electrical differential across the cell membrane. Empirical.
 * Equilibrium potential = calculated from Nernst equation for a particular ion. __Notice that the equilibrium potential of all permeating, non-pumped ions have to be equal to the membrane potential, at equilibrium.__
 * On membrane potential: by convention, notated about the inside of the cell with respect to the outside of the cell (so a negative membrane potential means there's a potential for positive ions to flow into the cell).
 * Know that each and every ion species has its own, independent equilibrium potential:
 * The equilibrium potential is dependent on particular electrical properties of the ion.
 * However, as above, note that for a non-pumped, permeating ion species (ie. Na+ or Cl-), the equilibrium potential is equal to the membrane potential for that membrane.
 * Understand that the number of excess anions in a typical cell is small compared to the total number of anions:
 * Doesn't need to be big to keep distinct ion concentration gradients in place.
 * Know that bulk solutions are always electrically neutral:
 * This, I think, is effectively because electrostatic repulsion doesn't allow charged solutions to stay together (see Betz's analogy about a one-liter box of cations needing a quarter of the weight of the earth to keep the lid on).
 * Technically this is not true across cell membranes - there's a small electric differential of ~-60 millivolts - but the electrical difference is small enough that we can get away with it.
 * What this means: in any given bulk fluid (ie. ECF/ICF) the electric charges must equal out.
 * This is able to keep concentration gradients away from equilibrium- that is, being electrically neutral takes precedence over being at a state of concentration equilibrium.
 * The cell is able to take advantage of this fact to maintain different compositions of ions inside and outside the cell.
 * Understand how an artificial cell can be in a state of equilibrium even though the concentrations of ions are not the same inside and out:
 * Okay. It's in a state of equilibrium because the electrostatic forces keep the ions from sliding along their concentration gradients.
 * Basically you've balanced the electrical equilibrium with the concentration equilibrium, This means that the cell isn't electrically neutral (though it's very close), and it also means that the cell isn't at a concentration neutrality (ie. different species of ions have different concentrations across the membrane).
 * Not to wax philosophical, but this is what allows us to be alive. Maintaining differences between things in the face of the 2nd law of thermodynamics - which drives things to a universal diffusion as the greatest expression of entropy - is what allows us to control fluxes in those differences, which in turn is the basis for pretty much all life as we know it, from arterial gas exchange to action potentials to which football team you root for.
 * Notice this electrical-vs.-concentration gradient thing does __not__ apply to uncharged solute atoms (electrostatic forces don't have any grip on uncharged atoms).
 * Be able to apply osmotic balance, charge neutrality, and the Nernst equation to calculate ion concentrations and the membrane potential of an artificial cell:
 * Nernst equation: Equilibrium potential is proportional to the natural log of the concentration of a given ion outside the cell over its concentration inside the cell.
 * ie: E is proportional to ln ([Iono]/[Ioni]) divided by the valence of the ion (+1 for Na+, -1 for Cl-, etc), where [Iono] is the concentration of the ion outside.
 * Approximation: E = 60/z * log ([Iono]/[Ioni]), where z is the valence (+1 for Na+, -1 for Cl-, etc).
 * (important) __Donnan equation__: [Ko][Clo] = [Ki][Cli]
 * Notes on possible test questions:
 * Dr. Betz seems to be fond of making charts where, given known and unknown concentrations of various ions inside and outside the cell, you fill in the blanks. There's a three-part process to solving these. This is what you need, not in any particular order:
 * (1) B__alance the electric charges //in each compartment//__ (ECF + ICF). ECF and ICF are always electrically neutral, more or less. So if inside a cell you've got 200 mosM of Na+ (valence +1) and 100 mosM of K+ (valence +1) and 50 mosM of proteins with a valence of -1, the total charge inside the cell right now is ((200*1) + (100*1) + (50*-1) = 250). To make it neutral, then, you need 250 mosM of Cl- (250*-1). If the valence of the proteins was -2, then the total charge inside the cell is ((200*1) + (150*1) + (50*-2) = 200). In which case you'd only need 200 mosM of Cl- to make it neutral. If this doesn't make sense, go find someone who understands it and threaten them until they verbally explain it.
 * (2) __Balance the osmolarity //between compartments//__ . If you add all the osmolarity of the ions inside the cell, it needs to be the same as the total osmolarity of the ions outside the cell. So if you've got 200 mosM of Na+, 100 mosM of K+, 50 mosM of proteins, and 250 mosM of Cl- in the ICF, total osmolarity = (200 + 100 + 50 + 250 = 600 mosM). This means that the total osmolarity in the ECF also has to be 600 mosM. So if you're missing one concentration in the ECF (in this example), just total the rest of them and subtract them from 600; what's less is what you have to have to balance the osmolarity. Again, if this doesn't make sense to you, find someone what can verbally explain it.
 * (3) __Use Donnan equation to get [K+] and [Cl-] for both compartments__. Because the product of [K+] and [Cl-] in the ICF is the same as the product of the concentrations of those ions outside the cell, if you know three of the concentrations, you can find the fourth. Ie: if AB = CD, and you know A, B, and C, D is pretty easy assuming you passed 9th grade algebra (which, for the record, I nearly didn't).
 * Something else he spent some time on in class: given a permeating ion's equilibrium potential (E) and the membrane potential (Vm) of the membrane, can you tell if there's an ion pump operating, and if so, in which direction it's operating?
 * This first part is easy. If a permeating ion's E isn't the same as the Vm, then there's a pump at work.
 * To figure out which direction it's going in, compare E and Vm.
 * If E is larger (more positive) than Vm, the ion wants to make the inside of the cell __more positive__.
 * This means if the ion's positively charged, it wants to go into the cell. Conversely, if the ion's negatively charged, it wants to go out of the cell.
 * If E is smaller (more negative) than Vm, the ion wants to make the inside of the cell __more negative__.
 * This means if the ion's positively charged, it wants to go out of the cell. Conversely, if the ion's negatively charged, it wants to go into the cell.
 * Try this on K and Na: K's E = -80, Na's E = +60. An average membrane potential is about -60.
 * Na's E is more positive than Vm, and it's a positively charged ion, so it wants to go into the cell.
 * K's E is more negative than Vm, and it's a positively charged ion, so it wants to go out of the cell.
 * Once you've figured out which direction the ion __wants__ to go, flip it. Then you know which direction the pump has to be pumping it to stop the leak in its preferred direction.
 * This is kind of a cheat sheet. But it's good to work through it on your own as well if you like mind pain.
 * Notice that, given the concentrations of ions inside and outside the cell, you can figure out E for those ions. So really, all you need to solve these problems is Vm and the concentrations of the ions you're looking at.

Other, random, notes: make membrane permeable to chloride but not sodium. Chloride atoms come into the cell, bringing a negative charge- when enough Cl ions are in there, electrostatic repulsion. Then when any Cl ion comes in along its concentration gradient, one gets kicked out by electrostatic repulsion. For K: same idea. Electrical gradient wants to keep K inside cell (since cell is negatively charged), even though concentration gradient wants to take K out of cell.

Na pump: transports Na out, K in.
 * In a real cell: the equilibrium potential for sodium is about +60. The equilibrium potential for potassium is about -80. The actual membrane potential is about -60. This means, since both K and Na are permeating, that there's a constant leak of K out of the cell (to make it more negative to approach equil potential) and a constant leak of Na into the cell (to make it more positive to approach equil potential). Thus rely on pump to pull back towards -60 as Vm. In other words, the current carried by sodium into the cell must equal the current carried by potassium out of the cell at equilibrium.

If you allow Na to come into cell (ie if you make the membrane more permeable to Na), get depolarization- cell membrane potential rises towards 60 mV.
 * Take-home here: any time you increase the membrane permeability of one ion, the membrane potential of the cell moves toward the equilibrium potential of that ion. And recall that the concentration gradient across the membrane determines the equilibrium potential for that ion. "The ion with the highest permeability wins."

Put another way: the ratio of the permeability of sodium to the permeability of potassium in a given cell is what determines the membrane potential of that cell. Notice that the permeability of a given membrane to an ion is largely determined by the number of open channels for that ion in that membrane. So if you have more open K channels than Na channels, your membrane is effectively more permeable to K than Na (and thus membrane potential will be closer to the equilibrium potential for K, -80, than it will be to the E of Na, 60).

=Acids, Bases, Buffers=
 * **Describe the law of mass action [le Chatelier's principle]:**
 * For any reversible reaction, the forward and reverse rates of the reaction are directly dependent on the concentrations of the __reactants__ and the __products__ // respectively //.
 * Ie: in A <--> B, if you increase the concentration of A, the rate of A -> B goes up. If you increase the concentration of B, the rate of B -> A goes up instead.
 * Equilibrium constant for a reaction Keq = [products] / [reactants].
 * Keq of water = 1.8 x 10-16 = [H+][OH-] / [H2O].
 * [H+][OH-] = 1 x 10-14.
 * This is universally, always, everywhere, true. No exceptions. Even for you.
 * In a neutral solution, [H+] = [OH-] = 1 x 10-7.


 * Define pH and pKa:

pH is the negative log of the proton concentration in a solution.
 * ie: pH = -log [H+].
 * In neutral solutions, [H+] = 1 x 10-7; log [H+] = -7, negative log = 7. **
 * That is: in neutral solutions, pH = 7, which is what you already know.

Side notes: histidine is the only amino acid side chain that protonates or deprotonates at physiological pHs- thus important for acid-base balance in body.
 * Notice this means that small differences in pH mean large differences in the concentration of protons-- which our bodies generally handle very poorly. **
 * Ka measures the tendency of a weak acid to dissociate in water.
 * pKa ** is the negative log of Ka for a given weak acid. **
 * The lower the pKa, the stronger the acid (more proton donation in water).
 * The higher the pKa, the weaker the acid (more proton acceptance in water).
 * **Write the Henderson-Hasselbalch (H-H) equation for any given weak acid or base:**
 * pH = pKa + log ([A-]/[HA])** . **
 * This means: pH = pKa + log ([proton acceptor]/[protein donor]).
 * Which means that, given the ratio of dissociated to undissociated acids in solution, you can calculate the pH of the system given the pKa, or can calculate pKa given the pH, or if you're given the pH and the pKa, can calculate the ratio of undissociated to dissociated acid in a given solution.
 * (ie: given two elements of that equation can get the third.)
 * Notice that when pH = pKa, the concentrations of deprotonated and protonated acid are equal.
 * (ie: when pH = pKa, the acid is half deprotonated and half protonated.)
 * **Define the H-H equation for the bicarbonate buffer system in extracellular fluid and use it to evaluate clinical lab data:**
 * pH = 6.1 + log ([HCO3-]/(0.3*PCO2))** . **
 * Essentially: the deprotonated bicarbonate (HCO3-) levels inversely vary with the partial pressure [levels] of CO2 in the blood.
 * What the hell that means: given the partial pressure of CO2 in the blood and the pH of the blood, can calculate the concentration of bicarbonate; or given the partial pressure of CO2 and the concentration of bicarbonate, can calculate the pH of the blood; or given the concentration of bicarbonate and the pH, can calculate the partial pressure of CO2 in the blood.
 * (ie: given two elements of that equation can get the third.)
 * **Important note: [HCO3-] needs to be measured in** millimolar** units; PCO2 needs to be measured in **millimeters mercury ** or **mm Hg** . **
 * Some background on all this mess: the buffering system (see below for details on buffering) in the body is predominantly made up of carbonic acid-to-bicarbonate: H2CO3 <--> H+ + HCO3-
 * This depends on the carbon dioxide levels in the blood-- dissolved CO2 reacts with water to form H2CO3.
 * This means that the equilibrium of the dissociation of H2CO3 is driven by the levels of CO2 in the blood (by le Chatelier's principle).
 * This works as a buffer, even though the nominal pKa of H2CO3 is 3.8 (thus would buffer optimally at 2.8-4.8, way outside physiological pH), because ion exchange in the kidney and input of CO2 changes the effective, actual pKa.
 * Define normal blood pH, [HCO3-] and pCO2 (please see your handout or learning objective):
 * Normal arterial blood pH: 7.34-7.44
 * Normal venous blood ph: 7.28-7.42 (lower due to higher [CO2])
 * Normal concentration of HCO3- in blood: 24 mM
 * Normal PCO2 in blood: 40 mm Hg
 * Normal concentration of CO2 in blood (he didn't ask, but it's in his practice problems): 1.2 mM
 * Describe how weak acids and bases work to buffer pH and define the pH range of maximal buffering capacity:
 * Effectively, you have a mix of weak bases and acids: the weak acids neutralize added bases without changing the pH much, the weak bases neutralize added acids without changing the pH much.
 * Optimal buffering capacity is determined by the pKa of the buffering acids/bases. There's a whole lot of titration chemistry behind this. Effectively there's a pH at which your buffer system can neutralize added acids or bases with very little change in overall pH, which is what you want to avoid a lot of big jumps in your blood pH (which would kill you): __this optimal buffering pH is plus or minus 1 pH unit from the pKa of the acid/base__.
 * Use the H-H equation to solve problems of how pH changes in defined buffers i.e. you should be able to determine how many equivalents of acid or base are needed to titrate the ionizable groups(s) of a weak acid or base from a starting pH to a final pH, given its concentration and pKa:
 * Quick way to think about this: every factor of ten that you have of deprotonated acid (A-) more than protonated acid (HA), you raise the pH by 1 over the pKa. If you have 1 HA molecule and 10 A- molecules, you just raised the pH by 1 from the pKa. If you have 1 HA and 100 A-, now the pH will be 2 plus the pKa.
 * Conversely, every factor of ten that you have of HA more than A-, you lower the pH by 1 from the pKa. So if you've got 10 HA molecules and 1 A- molecule, the pH of the solution is going to be the pKa minus 1.
 * So if you start out with a pH of 6 and the pKa is 6, to get to a pH of 7, you'll need to add in ten times the current amount of A- (which will add one to the pKa and result in a pH of 7). If you want to get to pH 5, you need to add in ten times the current amount of HA. pH 8: 100 times the current concentration of A-. pH 4: 100 times the current concentration of HA. Etc.
 * Again: find someone who gets this and beat or plead an explanation out of them if this is Greek to you.

Notice that the kidney controls a lot of pH by retaining or excreting HCO3- and NH4+.

Notice that the low pH in the stomach, which allows compounds to be easily protonated, allows drugs that need to be protonated to be absorbed into the cell to be so-

Remember: Higher levels of CO2 in the blood lower the blood's pH. This is because you make more carbonic acid, which drives the deprotonization of carbonic acid to form bicarbonate + a proton. These extra protons float around in the blood and lower the pH.

=Membrane Structure= has another order of magnitude of surface area over the intracellular membrane area. o Notice that under physiological pH, lipid bilayers spontaneously reform compartments to minimize contact of their hydrophobic tails with water.
 * Describe the lipid components of biological membranes and what differences exist between membranes of the different organelles
 * Lipid bilayers: roughly 5 nm thick, spanned by proteins.
 * Head groups: polar. Tail groups (inside the membrane): hydrophobic.
 * Classes of lipids in membranes:
 * Phosphoglycerides
 * Phosphotidylethanolamine
 * Phosphotidylcholine
 * Phosphotidylserine
 * Phosphotidylinositol
 * Sphingolipids
 * Cholesterol
 * All are synthesized at least in part in the ER and have polar and nonpolar ends.
 * __Phosphoglycerides__: have a glycerol-3-phosphate backbone with 2 fatty acyl chains (either saturated or mono- or polyunsaturated) at one end and a polar head group at the other end. What type of head group determines which phosphatidylglycerol it is (ethanolamine, inositol, etc).
 * __Sphingolipids__: have a 'sphingosine' (long acyl unit) with either of two polar head groups (which one it is distinguishes which type of sphingolipid it is) and an attached fatty acid chain.
 * __Sphingomyelin__: phosphocholine head group.
 * These are started to be made in the ER and finished in the Golgi apparatus.
 * __Cholesterol__: have a characteristic hydroxylated ring structure at one end, and a fatty acid chain on the other.
 * About 30% of all genomic proteins are membrane proteins
 * There are some double-membrane (mitochondria) and some single-membrane (nucleus) organelles.
 * Notice that there's roughly ten times the surface area of the plasma membrane contained as surface area of membranes within the cell; notice that the cytoskeleton
 * Bilayers are asymmetric. Notice that particular phospholipids don't flip from one side of the membrane to another without enzyme activity, but lots of lateral diffusion all the time.
 * [Not on LO's so learn at own risk:]
 * [Exoplasmic face: phosphotidylcholine, sphingomyelin, glycolipids (glycosylated lipids).
 * Cytoplasmic face: phosphoidylethanolamine, phosphatidylserine, phosphatidylinositol.]
 * Cholesterol is probably roughly equally distributed on both sides of the membrane.
 * Understand how concept of membrane fluidity and how membrane fluidity is modified
 * Membrane fluidity: the degree to which lateral motion is possible among adjacent phospholipids on a given side of the bilayer. Some lipids are anchored, some are free-floating within any given membrane.
 * Notice that different physical compartments of a membrane can have different degrees of fluidity.
 * Having acyl chains unsaturated makes the membrane more fluid (less tightly packed).
 * Cholesterol, when intercalated in membranes, stiffens the membrane and makes it less fluid.
 * Membrane composition determines thickness and curvature:
 * Membranes high in cholesterol also thicken the membrane by straightening out the nonpolar, hydrophobic groups inside the bilayer.
 * Curvature is dependent on the size of the polar head groups on both sides of the membranes.
 * Understand the many different ways proteins interact with membranes
 * [Cholesterol production: Proteins SCAP and SREBP, under high-cholesterol conditions, stay inactive in the ER. Under low-cholesterol conditions, SCAP and SREBP go on to the Golgi and wind up triggering nuclear receptors to synthesize more cholesterol.]
 * Different kinds of membrane proteins:
 * __Integral__ proteins (fully or partially embedded in membrane)-- can go multiple times back and forth through the membrane (multiple transmembrane domains).
 * Notice that transmembrane domains are usually alpha-helices, but can also have beta-sheet barrel formations (usually bacterial).
 * Can have transmembrane proteins that are only bound to one side of the bilayer.
 * Can have proteins that are only attached at their ends to the membranes.
 * __Peripheral__ membrane proteins: covalently interacting with proteins bound in the membrane, but not themselves embedded in the membrane.
 * Describe the importance of sphingolipids and cholesterol in biological membranes
 * Cholesterol: see above (thickness and nonfluidity).
 * Lipid rafts: formed primarily from sphingomyelin and cholesterol, thus extremely nonfluid-- effectively a lipid platform for various proteins. Used for signaling processes.
 * Although the different carriers and channels that allow molecules to be transported across membranes is discussed by Dr. Betz in his lectures, understand how these molecules' function should be integrated into the function of biological membranes.

=Membrane Fusion=
 * Understand the importance of sub-cellular protein targeting.
 * A cell has a lot of compartments through which it needs to move big proteins without compromising the membranes' integrity-- thus transport vesicles (membrane package-wrap, made from membrane of membrane-bound organelle of origin, containing cargo to be transported to target membrane-bound organelles).
 * This (vesicular transport) is the basic principle of transporting substances from one intracellular compartment to another.
 * Incorrect transport frequently results in disease: cystic fibrosis is due to a failure in chloride ion channel-protein transport.
 * Myelin/tubulin cytoskeleton elements: act as "highways" to transport vesicles throughout cell. "Motor" proteins use ATP to drive vesicles along these highways (actin for myelin, kinesin/dynein for tubulin). The particular motor proteins are specific for particular destinations-- one variety of kinesin may take a protein to one location, another to another, etc.
 * Know the basic principles of membrane and viral fusion:
 * Membranes don't just automatically merge when they get close together (which is good, otherwise things would not be well separated in the cell).
 * Two membranes: begin with them coated with water molecules. First you have to remove the water before you can fuse the lipids; then you need to overcome charge repulsion between lipids in order to allow a greater efficiency of merging. Also need to make sure these specific lipids should be merging in the first place (specificity of membrane fusion).
 * Early research: Botulin and tetanus toxins inhibit vesicle secretion by neurotransmitters. They do this by cleaving particular proteins: the SNARE proteins.
 * Know the function and structure of SNARE proteins:
 * Come in three shapes:
 * VAMPs: transmembrane domain at one end, with a helical domain in it.
 * Syntaxin: transmembrane domain, still a helical domain.
 * SNAP25: no transmembrane domain, two helical domains, fatty acid binding region that more or less acts as a membrane-binding domain.
 * (notice that there are different flavors of each protein, each of which can only bind to certain flavors of their counterparts.)
 * The helical domains of all three proteins serve to bind with each other-- form an aligned bundle or tetramer.
 * All __vesicles__ contain VAMPs. All __target membranes__ contain syntaxin and SNAP25.
 * When the vesicle nears the target membrane, the helical domains on the vesicle bind to those on the target membrane: these effectively press the two lipid layers together, squeeze out water and overcome charge repulsion, and promote the lipid fusion.
 * To do this, the helical binding needs to be extremely strong, and so it is.
 * This means a problem after the lipids have fused: how do you get apart the SNARE proteins once they've bound together so strongly?
 * Know regulation of SNARE-based fusion:
 * Enzyme that regulates the dissociation of SNARE proteins: //NSF// protein. Forms a kind of turning barrel around the SNARE complex and twists it apart with the help of a lot of ATP.
 * After being unwound, syntaxin becomes unfolded; need another protein, //Sec1//, to refold it properly.
 * __SNARE cycle__: VAMP on vesicle, syntaxin and SNAP25 on target membrane, helical binding and lipid fusion, NSF-mediated unwinding, refolding of syntaxin with Sec1.
 * Know the mechanism of viral fusion:
 * ["__Envelope viruses__" (influenza, ebola, HIV, etc): unlike other viruses, it has a membrane envelope around its capsid (capsid: protein structure containing genetic material).]
 * In order to infect a cell, the envelope needs to fuse with the target membrane.
 * Notice that the target isn't always the plasma membrane: influenza virus gets endocytosed first, then when it's inside the cell it goes after the endosome membrane.
 * Notice that when a cell becomes infected by an envelope virus, the cell makes viral fusogenic proteins that go to the cell's surface-- will put these fusogenic proteins on emergent viral capsids.
 * Mechanism is similar to SNARE-mediated fusion, but virus has only one protein:
 * Has a transmembrane domain as well as a non-transmembrane hydrophobic domain hidden inside the folded protein.
 * Binding to the cell surface causes the exposure of this hidden hydrophobic domain, which inserts into the host membrane.
 * The virus has a lot of these proteins on its envelope: effectively they bind the envelope to the host membrane.
 * The other feature of the protein is multiple helical domains: one they're bound to the host membrane, these domains bind strongly to each other and ratchet the envelope tightly onto the host membrane, squeezing out water and overcoming charge repulsion as per the SNARE mechanism.
 * Know the regulation of viral fusion:
 * Only thing I can think of that he mentioned was that influenza viruses are dependent on a cellular drop in pH to release their genetic material into the cell.

=Membrane Potential III=
 * Know the difference between equilibrium and steady state:
 * Equilibrium: Ion concentrations are not changing, inside or out, over time.
 * Steady-state: Ion concentrations are not changing, inside or out, over time because the cell is furiously working to keep them that way.
 * Equilibrium is Zen (no ions have any striving ideas to go in or come out).
 * Steady-state is a contained prison riot occurring at the same time as an attempted storming of the Bastille (lots of things want to go out, lots of things want to come in, the guards are beating everyone the hell up to keep them put).
 * In a cell at rest (steady state), given an ion's concentration inside and out, the membrane potential, and knowledge that the membrane is permeable to the ion in question, be able to determine whether or not a pump for that ion must exist, and if so, which direction it pumps the ion:
 * I went over this in detail at the end of the notes for "Membrane Potential II".
 * Understand that membrane potential depends on relative, not absolute, permeabilities to ion:
 * Example: a membrane is very permeable to sodium but not to potassium. The membrane potential then approaches the equilibrium potential of sodium (+60). If the membrane is very permeable to potassium but not to sodium, the Vm approaches the E of potassium (-80). If they permeabilities are equal, the Vm won't change - within a first approximation - no matter how much absolute quantity there is (note, however, that if the membrane gets really really permeable, the Vm will indeed change and approach 0 as the changing concentrations of the ions make their Es approach 0).
 * Understand that the short term determinant of membrane potential is not the Na/K pump, but relative membrane permeabilities:
 * Depending on the size and the number of channels in the membranes (larger cells with more channels are more vulnerable, quicker, to Na/K pump failure than others).
 * Define Driving Force of an ion:
 * The ion "wants" to have the membrane potential approach its own equilibrium potential. It will drive the membrane potential towards that equilibrium, aided or counteracted by other ions, which are also driving the membrane potential towards their own equilibrium potentials.
 * Understand why, in neurons and other excitable cells, membrane potential is sensitive to small changes in [K+]o, but not [Na+]o:
 * (Understand that [Na+o] is the sodium concentration outside the cell.)
 * (1) Because the membrane potential of the cell is nearly at the E of K+ anyway, the loss of extracellular Na+ (which brings the E of Na+ closer to 0) will make the Vm slightly hyperpolarized (closer to -80 mV) but have not much other effect.
 * (2) [K+]o is more sensitive because its starting concentration outside the cell are much smaller than Na+, and thus is much more sensitive to small changes in concentration.
 * Know the mechanisms by which calcium, glucose + insulin, Na/K ion exchange resins, and renal dialysis counteract the signs of hyperkalemia:
 * Hyperkalemia: Excess of potassium in blood (outside cells, in ECF). This changes the membrane potential of the cell because it alters the concentration gradient on which the electrical gradient of the membrane is based.
 * Effectively, this raises the equilibrium potential of K+, which in turn makes the membrane potential rise substantially (by ~20 mV for a 2% calcium efflux).
 * This is almost universally fatal. The reason it's so dangerous is that messing with membrane potentials is a good way to screw up action potential propagation, particularly in the pacemakers in the heart-- thus can't get heart contraction signals.
 * Caused by a variety of things: massively lysed cells, kidney failure, severe muscle damage.
 * Treatment:
 * Can stabilize cardiac conduction system by giving Ca2+
 * Can get cells to take up the K+ by giving insulin and glucose to stimulate Na-K pump. (Insulin gets glucose into the cell, where it activates the pump to work harder).
 * If you give alka-seltzer and alkylyze blood, can stimulate proton pumps to get K+ into cells.
 * Can use cation exchange resins: Effectively, Na+-lined "flypaper" for K+ in the blood, swapping out Na+ for K+.
 * Can use renal dialysis to pherese excess K+ out of blood.

=Membrane Transport=
 * [Transporters: Proteins in the membrane that move specific substances, as in moving particular large molecule or pumping ions against their gradients.
 * Know the difference between primary and secondary active transport:
 * Primary active transporters: use ATP for energy source.
 * Ie: Na+-K+ pump, Ca2+ pump, proton pumps.
 * Secondary active transporters: don't use ATP. Uses energy derived indirectly from primary pumps.
 * Amino acid transporters: intake AA into cells; derives energy from leakage of Na+ into cell (slight amount of energy released by Na+ leak).
 * H+-to-Na+ exchange (protons out powered by energy of Na coming in).
 * Define cotransport and exchange transport:
 * Cotransport: secondary active transporters that move different solute species in the same direction.
 * Exchange transport: secondary active transporters that move solutes in opposite directions.
 * Na-Ca pump: Na+ in cell, Ca2+ out of cell (or, sometimes, the other direction)
 * A note about calcium: [Ca2+] inside the cell is kept extremely low (about 0.1 micromolar; about 10,000 times lower than outside the cell).
 * Thus: calcium has very high equilibrium potential (~120 mV). This allows single-channel openings to allow Ca2+ in can have large effects inside the cell.
 * The Na-Ca exchanger effectively works by balancing the favorable energy from 3Na+ entering the cell with the unfavorable energy from Ca2+ exiting the cell.
 * Notice that this can change direction depending on the current state of the membrane potential and the concentrations of Ca and K in the cell.
 * Be able to determine which direction a non-electrogenic secondary active transporter will operate, given the concentrations of the participating ions inside and out:
 * Electrogenic: one cycle produces a net charge transfer across the membrane. Thus non-electrogenic are transporters that don't change the net charge across the membrane. In this case, my guess would be that they tend to follow the direction of ion concentration (see the theoretical H/K exchanger below).
 * Know that electrogenic secondary active transporters can reverse direction, depending on membrane potential.
 * (see under Na-Ca pump above)
 * Understand the non-existence, but conceptually useful idea, of the H/K exchanger:
 * If you drop the proton concentration in the blood, can promote cell uptake of K in exchange for protons going out into the blood, and vice versa. The H-K exchanger doesn't actually exist as an entity, but you can think about it like that.
 * Essentially: if you have excess K, you can give bicarbonate (HCO3-) to decrease the total H+ concentration in the blood. When that happens, the exchanger that switches K for H swaps out K in the blood for H in the cell, thus making the proton concentration in the cell go down and the potassium concentration in the cell go up. This effectively takes K out of the blood, treating hyperkalemia.
 * Know how cells can concentrate glucose inside, even though glucose transporter cannot pump glucose against its concentration gradient
 * Inside cell, glucose gets phosphorylated, which makes it unable to fit into the glucose transporter-- thus glucose flow is more or less unidirectional into the cell. Notice that glucose transporters are normally sequestered within vesicles in the cell until insulin signals the cell to make those vesicles merge with the cell surface and transport glucose into the cell.
 * Notice that the CHF drug Digitalis blocks the Na-K pump. It does this in order to increase the contractile strength of the heart muscle; what it does is raise the effective concentration of Na+ in the cell-- which then renders the Na-Ca pump less effective, allowing more Ca2+ to stay in the cell, increasing the strength of the muscle cell contraction.

=Epithelial Transport=
 * Epithelial cells: cells which sequester 'third space' fluids from extracellular fluid.
 * These cells are __polarized__: not electrically, but meaning that they transport in only one direction (apical to basolateral).
 * Side of epithelial cells facing the third-space fluid is called the '__apical__' or mucosal membrane; the side facing the ECF is the __basolateral__ or serosal membrane.
 * The basolateral membrane is generic across the entire surface.
 * The apical membrane is specialized and compartmentalized (with tight junctions) for special functions.
 * Mainly, these involve transporters of various kinds.
 * Tight junctions: here, protein-based 'glue' that holds nearby cells together but blocks lipid exchange with the basolateral membrane.
 * Understand the generic transport mechanisms for NaCl and water into the blood:
 * NaCl: Na+ picked up by the apical membrane (high permeability of apical membrane for Na+) and is transported into the cell, then is pumped out by the Na/K pump out the basolateral membrane. Cl- passively follows the Na+ to equalize the electrical potential.
 * Water follows the influx of NaCl into cells to balance osmolarity.
 * Notice all this is passive (no ATP usage) absorption.
 * Know the basic transport mechanisms by which glucose and amino acids are absorbed into the blood:
 * Epithelium in GI tract: AA, sugars, and glucose are pumped (secondary active transport) through the apical membrane and diffuse out the basolateral side passively into the blood.
 * AA and sugars are picked up by their target cells by secondary active transport mechanisms.
 * Glucose, as mentioned before, diffuses into cell along its gradient (but is phosphorylated in the cell to prevent efflux).
 * Know the major differences between 'tight' and 'leaky' epithelia:
 * Tight: more junction proteins, tighter seal between cells. Used particularly in the distal tubules of the kidney.
 * Loose: less junction proteins, looser seal between cells.
 * Tight: "fine tuning"-- strictly controlled substance transport at lower levels, don't want backflux.
 * Loose: quantity over quality (needs lots of transporters, not too picky about equal amounts).
 * Know that four important substances - water, O2, CO2, and urea are never pumped, but always move passively down their concentration gradients:
 * You can open and close aquaporin channels to allow the water to come in or go out, but it always flows along its gradient.
 * Understand the relative roles of the G.I. tract (minimal) and kidney (extensive) in excreting non-volatile metabolic wastes and regulating ECF composition:
 * GI tract excretes (in feces) largely things you couldn't absorb in the first place. GI tract absorbs pretty much everything it can that comes in to it indiscriminately (thus its small role in excreting waste).
 * However, GI tract does excrete dead red blood cells (excreted into the GI tract from the liver), which is significant.
 * Kidney concentrates waste very well in the urine- gets rid of approximately 0.5 moles of non-volatile (nongaseous) metabolic waste a day, mainly end-products of nitrogen metabolism (urea) and protons.
 * Given two of the following, be able to calculate the third: apical membrane potential, basolateral membrane potential, trans-epithelial potential:
 * Trans-epithelial membrane potential = basolateral membrane potential - apical membrane potential.
 * Notice trans-epithelial is abbreviated PD.
 * One thing to remember here is that the membrane potentials of the apical and the basolateral membrane will be different (different ion permeabilities, etc).
 * The other thing to remember is that membrane potentials are always written in the form of describing the inside of the membrane with respect to the outside-- ie, in a membrane potential is -10 mV, the inside of the cell is 10 mV more negative than the outside.
 * Know the basic process by which some epithelial cells secrete (rather than absorb) fluid.
 * There's a chloride-selective channel in the apical membrane in some epithelial cells, cAMP-gated on the inside of the cell to open and allow the efflux of Cl- back out of the apical membrane (which takes Na+ and water with it).
 * This process is driven by receptors on the basolateral side of the epithelial membrane that are triggered by acetylcholine to open those gates.
 * The reason the Cl- channels excrete Cl- instead of intaking it is that there's an non-electrogenic pump on the basolateral side that uses Na+ leakage to import Cl- into the cell.
 * Effectively you're putting watery/serous substance out into the epithelial secretions (mucus, etc).
 * This is the basis for cystic fibrosis: the chloride channel isn't properly implanted in the cell membrane, and thus no dilution of mucus secretions are possible.
 * This is also the basis for cholera: the cholera toxin gets into the cell and opens the Cl- channel without regulation- which leads to a massive efflux of water out the apical membrane into the epithelial system (diarrhea, etc).
 * Understand the main routes of excretion of metabolic wastes - CO2 and urea, in particular.
 * CO2, unsurprisingly, is excreted in the alveoli of the lungs.
 * Urea is excreted in the urine after being picked up out of the blood in the kidneys.

=Action Potential I=
 * Understand how the passive electrical properties of axons render them poor conductors of electrical signals over distances greater than a few millimeters:
 * The electrical resistance in cytoplasm is high- also the electrical insulation provided by the cell membrane is poor. Unassisted passive electrical conductance would allow the signal to completely decay before it reached its target.
 * Describe the analogy between an electrical 'booster station' and an action potential:
 * Essentially, at various "stations" along the axon, a power source (the gradient of Na+, that is, potential chemical energy) 'boosts' the electrical signal back its original strength, based on the 'readings' of the signal strength by voltage-sensing receptors.
 * In axons this 'reading' is done by __voltage-gated sodium channels__.
 * Know how changes in membrane resistance, membrane capacitance, and internal (axial) resistance affect the passive spread of voltage along an axon:
 * Okay. The cell membrane acts as a capacitor between the ICF and the ECF. Notice that as soon as current reaches the ECF it is effectively lost-- it's no good for signal transmission.
 * The ion channels in the cell act as resistors that slow the flow of current from the ICF to the ECF.
 * The resistance of the cytoplasm in the axon itself acts as a resistor that slow the flow of current along the ICF (in the direction you want the impulse to go, namely down the axon).
 * What this means if you want an axon to propagate quickly:
 * (a) You want the capacitance to be low (high capacitance delays the flow of current down the axon, since it's the first place your current goes).
 * (b) You want there to be lots of resistance to current leaving the cell (thus closing the ion channels).
 * (c) You want as little resistance as possible to current traveling down the axon.
 * The way the cell does this is (a) wrap the axons in myelin, which effectively increases the thickness of the membrane and thus drops the capacitance, speeding up the electrical propagation; (b) close ion channels to pump up resistance to current leaving the cell; and (c) increase the diameter of the axon, which increases current flow and effectively lowers internal (along-the-axon) resistance.
 * If this doesn't make much sense to you, you're in good company, or at least in mine. Just remember: thick membranes = low membrane capacitance = faster AP transmission. Fewer on channels = high membrane resistance = faster AP transmission. Larger-diameter axon = lower internal resistance = faster AP transmission.

=Action Potential II, III=
 * [Features of APs:]
 * (1) Propagated
 * (2) Vm reverses
 * (3) Brief (<1 ms)
 * (4) Threshold
 * (5) Refractory
 * (6) Accommodation
 * Describe the positions of the activation and inactivation gates in sodium channels during an action potential:
 * Sodium activation gates (m gates** ) are 'normally' closed at rest; they open (thus making the membrane more permeable to sodium) in response to a small depolarization (ie inside of the cell gets more positive).
 * Notice that the // m // gates are on the // outside // of the cell.


 * Sodium inactivation gates (**h gates** ) are 'normally' open at rest; they close (thus making the membrane less permeable to sodium) in response to the large depolarization that occurs after the //m// gates are opened. Notice there's a lag time between the point at which the //m// gates open and the point at which the //h// gates close (which allows the flood of Na+ to fully depolarize the cell to generate the AP). **


 * Notice that the //h// gates are on the //inside// of the cell.
 * Notice that there are also voltage-gated K+ channels that allow the cell to repolarize much faster after firing an AP (allow K+ out of cell to restore Vm after influx of Na+). These open around the time the AP signal is at the peak of its intensity (ie. when the cell is approaching maximum depolarization).


 * **Understand that intracellular concentrations of sodium and potassium do not change much after a single action potential:**

The flow in of Na+, and the flow out of K+, don't put much of a dent in the overall stores of both of those ions in the cell. Therefore an axon can fire multiple times before it needs to equilibrate its Na/K levels with the Na-K pump.


 * [Length constant λ = length along axon it takes for amplitude of electrical signal to decay to 37% of its original strength. In cellular axons this is about 1 mm.]
 * [Time constant = time it takes for an axon's signal to reach its maximum strength.]
 * Understand the role of the sodium/potassium pump during the action potential:
 * During the AP, the Na-K pump does pretty much nothing important. It's needed down the road to make sure that the Na-K concentrations are back to their proper levels (see accommodation below), but in the short term it's more or less irrelevant.
 * Describe the mechanisms underlying the refractory period of the action potential:
 * Recall that there are //two// refractory phases: the __absolute__, during which the cell will not fire an AP no matter what the stimulus, and the __relative__, during which the cell needs a much larger stimulus than normal to fire another AP.
 * This is due to the fact that the //h// gates (Na inactivation gates) require some time before they can reopen and allow Na into the cell (thus even if //m// gates open, the Na+ can't get into the cell).
 * During the absolute refractory phase, the //h// gates are more or less all shut.
 * During the relative refractory phase, the //h// gates are not all shut, but enough of them are that it will require more depolarizing stimulus to trigger the Na+ positive feedback cycle.
 * Notice that the K+ channels also contribute to the refractory period: it takes them a little while to close after repolarizing the cell, which help make the cell briefly hyperpolarized during the refractory period and makes it harder for the cell to become depolarized enough to trigger another AP.
 * Notice that the refractory period is the basis for the unidirectionality of an AP: as the impulse travels down an axon, it can't travel backwards because the portion of the axon just behind it (the one that just fired the AP) is in its refractory period.
 * (note: here I use "unidirectionality" to refer to the fact that an AP can't go backwards. Remember, thought, that if you start an AP in the middle of an axon, it goes in __both__ directions down the axons. One way of thinking about this is that __the direction of conduction down an axon__ isn't unidirectional, but __the direction a given AP travels__ is.)
 * **Describe the mechanisms underlying accommodation of the action potential:**
 * **Accommodation** : If a progressively larger current is applied to the cell slowly, often an AP won't fire-- if the depolarization is slow, the inactivation ( // h // ) gates have enough time to close before the AP has a chance to get going.
 * This is the basis for the problem with hyperkalemia: you wind up with a slow depolarization of the membrane that prevents APs from firing by slowly activating the // h // gates and preventing sudden depolarization by positive feedback.


 * This means that the more APs a cell fires, the more steadily depolarized it becomes. This, in turn, means that after a certain point of depolarization, the //h// gates (inactivation) are shut, and stay shut, meaning that the cell can't fire more APs until the Na-K pump restores its normal concentrations.

When this happens depends on the axon's diameter. Basically, the diameter of the axon determines its surface-area-to-volume ratio: the larger the axon, the smaller this ratio, which means that its Na-K balance is disturbed relatively less per AP than a smaller-diameter axon.
 * Thus: large-diameter axons accommodate more slowly; small-diameter axons accommodate more slowly.]


 * Define threshold for an action potential:
 * AP **threshold** is the point at which the incoming current of Na+ __equals__ the outgoing current of K+ (as Na+ comes in, K+ wants to come out more strongly). If the incoming current (Na+) gets just a little higher, the cell will enter the positive-feedback loop of depolarization.
 * A more intuitive way of thinking about it is that the threshold is the point at which just a hair more depolarization will probably make the cell fire an AP and just a hair less depolarization will probably make the cell return to normal polarization levels.
 * Understand the explosive, positive-feedback nature of the rising phase of the action potential:
 * If a cell is depolarized past its threshold point, the depolarization of the cell causes Na+ channels to open (//__m__// __gates__), which causes the cell to depolarize even faster, which causes more //m// gates to open, etc, etc.
 * Describe how action potential propagation relies on voltage-gated sodium channels acting like molecular 'booster stations'
 * See "Action Potential I." Effectively the passive propagation of the AP is sufficient to depolarize the next segment of axon enough to cause that second segment's //m// gates to open, causing another AP-- and so on down the axon.
 * [Basis for anesthestic: block //m// gates so that depolarization can't happen.]
 * [Notice: __Difference between small and large nerve fibers__:]
 * Small (ie pain and temperature fibers):
 * Slower AP conduction.
 * Have a high threshold to extracellular stimulation (harder to trigger AP).
 * Lower safety factor: easier to block //m// gates with anesthetic.
 * Large (ie touch receptor and positional fibers):
 * Faster AP conduction.
 * Have a lower threshold to extracellular stimulation (easier to trigger AP).
 * Higher safety factor: harder to block //m// gates with anesthetic.
 * Know why action potential propagation is much slower than the velocity of light:
 * Aside from the fact that it relies on the movement of ions (not massless particles), the fact that it's moving through a fluid with resistance as opposed to through a vacuum or through low-resistance copper wire, and the fact that it relies on a slow threshold-gated signal for propagation? No idea.
 * Know what myelination does to axonal membrane resistance and membrane capacitance, and why these changes increase conduction velocity
 * __Myelination__ effectively decreases the __capacitance__ of the axonal membrane (making a signal go __faster__, no waiting for the capacitance to build) and increases the membrane __resistance__ (making a signal go __farther__, no leaking of ions out of the membrane).
 * Describe refractoriness, and explain how it prevents an action potential from reversing its direction of propagation:
 * See above, under "refractory period."
 * Understand why demyelination can slow or block action potential conduction:
 * Well, if myelination allows the AP to go faster through increased capacitance, demyelination can make an AP go slower. Likewise, if myelination allows an AP to go farther through increased membrane resistance, demyelinated can make it go a smaller effective distance, which may not be far enough to reach the next node of Ranvier and set off the next AP.
 * Describe the effect of extracellular calcium ions on action potential threshold:
 * Ca2+ in the ECF normally remains bound to negative charges on the outer surface of the cell.
 * However, in hypocalcemia, [Ca2+] in the ECF goes down, which allows some of those negative charges to go unbound. This makes the outside of the membrane, at particular places where the Ca2+ is thin, selectively more negative.
 * As the outside of the membrane gets more negative in places, the potential difference across the membrane at those places goes lessens (ie, it depolarizes), thus triggering the //m// gates to create an AP without normal signaling.
 * Note the difference between hyperkalemia and hypocalcemia:
 * Hyperkalemia: excess of [K+] in ECF causes gradual depolarization of the entire membrane, which allows //h// gates to shut and stay shut, preventing APs from being generated.
 * Hypocalcemia: insufficiency of [Ca2+] in ECF caused localized depolarization of the //outside// of the membrane, which causes //m// gates (which are on the outside) to open and cause APs to be generated erratically.
 * [First sign of this is often "Trousseau's sign": contraction of the flexors in the elbows and wrists.]
 * Know the effect of axon diameter on conduction velocity, threshold to extracellular stimulation, safety factor for conduction, and likelihood of being myelinated
 * //Axons with a larger diameter conduct impulses more quickly.//
 * Interesting aside: notice that, given unlimited space, we'd want all of our axons to be huge-diameter to get really quick information to our brains. But given limited space, there's a tradeoff between //size// of axon (faster signal) and //number// of axons (more information in a slower signal). So some of our axons are really big (the get-out-of-the-way-of-the-bus axons) and some are smaller but more numerous (the ponder-the-meaning-of-life axons).
 * __Safety factor__: the fact that we have a lot more Na+ //m// gates than we really need to get to threshold and generate an AP of sufficient size in a given axon.
 * Means faster conduction is possible (depends on # //m// gates).
 * Also means can send APs down multiple branches of an axon.
 * Threshold to extracellular stimulation: As mentioned, large-diameter axons have a lower threshold to stimulation, while small-diameter axons have a higher threshold.
 * Smallest axons are unmyelinated: actually it winds up being more efficient this way.
 * Know the mechanisms by which calcium, glucose + insulin, Na/K ion exchange resins, and renal dialysis counteract the signs of hyperkalemia:
 * See "Membrane Potential III."