Rachel A. Kaplan and Elbert Heng Final Exam Review Rachel A. Kaplan and Elbert Heng

Announcements Your final is tomorrow; get hype! Things you should bring: A calculator Some pencils (or pens, if you want to be bold) Your brain!

What this review today will cover: As your exam is tomorrow, hopefully this isn’t the first time you’re going to be reviewing material accordingly, we will be: going over important topics and difficult concepts answering any questions you have providing moral support

Musings… You should study material that was tested on previous exams You should study material that was not tested on previous exams Most importantly: You should know the big important concepts that we’ve covered You should also spend time to review some details Look at our old slides for more comprehensive review In a nutshell: know everything. We’ll help you on which of the details you need to know (things that are in charts or we’ve told you to memorize in the past).

Be prepared to think… You have all the knowledge you need to answer these questions (all within the scope of the course) – just think hard and reason it out! The questions are designed to be answered either straight from knowledge or built from first principles.

Final Exam

THE BIG PICTURE: FIRST THIRD Fundamentals of synaptic transmission from an electrophysiological perspective Important Topics Ion Channels! Membrane Potentials - The Nernst Equation The Action Potential Membrane Properties Synaptic Transmission

THE BIG PICTURE: SECOND THIRD A little bit of everything, but mostly synaptic transmission from a molecular and cellular biological approach with electrophysiological implications Important Topics Vesicles: exo and endocytosis Indirect synaptic transmission Mechanosensation Behavioral Neurobiology Dendrites Electrical Synapses

THE BIG PICTURE: THIRD THIRD Plasticity and all its friends IMPORTANT TOPICS LTP and LTD Intrinsic Plasticity Learning Development LTP and Addiction The third paper On Kauer’s Lecture: review the slides

Ion Channels Ion channels pass ions This is studied with electrophysiological techniques Voltage Clamp Current Clamp (and putting these two together) I/V Plots Single Channel vs. Whole Cell Recording Single channels are constantly flickering open and shut; the population of channels will reflect the state of the cell Gating Modulation of Gating

Electrophysiology This is CURRENT CLAMP – clamp current (inject current) – measure change in voltage of membrane as response. LOOK AT THE AXES

Electrophysiology THIS IS VOLTAGE CLAMP – note the multiple voltages at which the membrane was clamped. The lines represent inward and outward currents measured as a response. This one doesn’t have axes, but you do have a clue to what’s going on as it shows the manipulations of membrane potential.

Electrophysiology I=Vg IV plots are extremely important to understand. They show channel characteristics. They let you see at x voltage the current will be y. MOST IMPORTANTLY: they show size or direction of current that flows at a given voltage. Which properties of the channel are shown in the plot? Reversal potential (+58mV here) – DETERMINED BY THE CONCENTRATION GRADIENTS OF IONS (NERNST EQUATION, COMING UP NEXT!) Conductance (slope of line) – conductance is a measure of how many ions a channel CAN PASS – distinct from how good a channel is at passing current. Therefore conductance depends on the ability of the channel to pass ions and the number of ions available for the channel to pass. This chart on this slide shows either a single channel’s IV plot or a non-voltage gated channel. What would a voltage gated channel’s IV curve look like? (See the next slide!) I=Vg

Electrophysiology This is a voltage gated channel’s IV plot. It shows that below a certain voltage (-60mV) it will not pass current in either direction. Let’s think about how an NMDAR’s I/V plot would look – it is a channel that is ligand gated (needs Glu) and voltage gated (needs depolarization to get rid of Mg++ block) It’s reversal potential would be different – because it’s permeable to both Na and K, (+58, -60mv respectively) it’s reversal potential (x-intercept would be at 0mV – the origin).

Gating and Modulation Gating: how the channel opens and closes S4 is the voltage sensor for VG channels e.g. Glu gates NMDARs and AMPARs Modulation: changes the open probability of the channel MODULATORS: Other subunits of the protein (beta subunits) Second messengers Changes in gene expression Phosphorylation Allosteric regulators Things that gate are required for channels to open Modulators don’t open channels

Nernst Equation vs. GHK Nernst: Single ion’s equilibrium potential. Equivalent to Vrev if a channel is singly selective for that ion. GHK: Combined equilibrium potential of all relevant (permeant) ions. Can give you the RMP Also can give you Vrev of multi-ion channels. So what could change the RMP? Altering ion concentrations! An example…. Decreasing Sodium Extracellularly – decrease inward current Decreasing Sodium Intracellularly – increase inward current

Membrane Properties All serve to modulate the speed of an action potential Membrane resistance (Rm) Membrane capacitance (Cm) Axial Resistance (Ra) Derived from these: Length Constant (λ) Time Constant (𝜏) All of the equations will be given to you if you would like to see the relationships written out… Membrane resistance – determined by leakyness- channel composition of membrane, presence of myelination Membrane capacitance – don’t worry about it too much, just think of it as a place where current goes before current goes through channels Axial resistance – an increase in axial resistance will decrease the current that can flow through the neurite (more resistant on the inside because few ions present, if the axon is thinner– has a smaller cross sectional area) Length constant – a longer length constant means that a given depolarization will cover a longer distance on the axon. Time constant – a larger time constant means that a given depolarization will travel for a longer period of time on the axon. Both of these are functions of capacitance – so the bigger the capacitance – the larger the two of these values both are.

Synaptic Transmission Llinas’ experiment Proved that calcium was necessary and sufficient for presynaptic transmitter release Depolarization is not sufficient! (if no calcium, no go) Quantal Hypothesis Quantum is a vesicle of neurotransmitter Quantal content - how many vesicles resleased! Quantal size – content of a single vesicle – how much NT is in it Content = mean EPP / average quantal sizeELBERT HERE

Mechanosensation Mechanosensitive neurons: Lots of receptor subtypes Generally: stretch-gated channels tethered to intra and extracellular matrices Fast, sensitive, adaptable (so that it can transduce a wide range of inputs), and specialized Lots of receptor subtypes E.g. Pacinian Corpuscles Respond to vibration because they are fast adapting Neuron is surrounded by epithelial cells that form many layers of gelatinous membranes called lamellae Pressure on causes neurons to fire Pressure off also causes neurons to fire In other words, they adapt

More Neurons/Proteins Involved Degenerin/ENaC Channels Respond to stretch/mechanical stimulation – slow adapting Meaning that they will stay open if they are continuously poked CEP Neuron Channels Senses viscosity of surrounding bacteria Rapidly adapting cation channels TRP-4: mechanosensory channel Other TRP Channels Sense temperature, chemical tastants TRP – transient receptor potential TRP channels were discovered to be channels because alterations in their pore sequence caused a change in the channel’s I/v curve – so if changing the i/v curve occurs with changing the protein, the protein is responsible for that i/v curve and therefore a channel.

TRP Channels Note the structure of the channel – single protein

Hearing and Proprioception Vibrations of air are transduced by mechanosensory hair cells Stereocilia are deflected, links between stereocilia are stretched, allows K+ inward current to depolarize cell Deflecting the other way will hyperpolarize the hair cell Stereocilia adapt by tightening tip links Movement of head in space is transduced by similar hair cells in other organs Utricle and sacculus – linear acceleration moves gel and crystals (otoliths), causes opening of hair cells Semicircular canals – rotational motion causes fluid in canals to move ampulla and embedded hair cells

Behavioral Neurobiology Responses to releasing stimuli e.g. Egg Rolling Stimulus (egg) triggers fixed action pattern e.g. Seagull Chick Feeding Stimulus (spot color) triggers pecking Supernormal stimuli: allows us to study nature of what an animal is actually responding to in a stimulus Releasing stimulus: a stimulus that triggers a stereotypic behavior Fixed action pattern: stereotypic, no end point for success, no check point during behavior, once initiated, must be executed in its entirety Supernormal stimuli: a stimulus that is bigger/better/sexier than the normal stimulus With stimuli, we can study the dimension of the releasing stimulus that actually is triggering the behavior. With egg: just that it looks like an egg, and how eggy it is (the bigger and egger it is, the more it triggers the behavior) With seagulls: REDness of the spot determines how good of stimulus it is, not how realistic the cardboard cut out of the parent is

Electrical Synapses Channels are composed of two Connexons Connexons are Hemichannels They are in turn composed of 6 connexins If all 6 connexins are the same protein: homomeric If different: heteromeric Most common connexons in the brain: Cx43 – Glial cells Cx36 – Brain neurons (perhaps the only connexon that is expressed in brain neurons!)

Electrical Synapse Physiology GJ provide high conductance pathway for ionic current to pass from one cell to another Ohmic (no voltage gating) Bidirectional Also pass small molecules like ATP, cyclic nucleotides So what would the electrophysiological recording of stimulation of a neuron that connected by a GJ to another neuron look like? Let’s take a closer look at the trace below… clearly shows spiking is bidirectional but greatly attenuated (diminished).

Gap Junction Evolution Pannexins / Innexins and Connexins are orthologues No sequence similarity but in teritiary structure are very similar Invertebrates do not express connexins Innexins and connexins can form GJs or functional hemichannels Pannexins only form hemichannels

Learning LTP and LTD are putative cellular mechanisms Shown with lots of experiments

Rabbits? The process of associative learning uses this circuit Input: sensory motor – tone- parallel fibers Also excites pons and deep nuclei directly (there are two pathways) Input: “error” signal – shock – climbing fibers Output: motor command- eye blink – purkinje cells LTD occurs in parallel fibers which means less inhibition of deep nuclei from purkinje cells Easier to express blinking behavior!

Know this pathway! Memorize the circuit and the nature of each connection. Stimulating the parallel fibers would be a substitute for the tone. Do this with other fibers! (e.g. inferior olive, purkinje cell).

Lashley Searched for the engram Equipotentiality Mass Action Equipotentiality: All parts of the cortex contribute equally to learning; one part can substitute for another part. Mass Action: The cortex works as a whole; performance improves when more of the cortex is involved.

Development of Circuits is: Activity Independent Sperry: chemoaffinity hypothesis Experiments Eye rotation Retinal ablation Stripe assay Mechanism Ephrins and Eph Receptors Activity Dependent Hebb: correlation based change Experiments Rewiring of A1/V1 Mechanism Synapse maturation LTP Depolarizing GABA Activity dependent gene expression

A little bit of both… Ocular dominance columns start to develop before eye opening but require activity to segregate more completely Spontaneous retinal waves may be responsible Ocular dominance shift: Monocularly deprived animals develop ocular dominance stripes but the open eye’s stripes are much wider

Putting it all together: Neural development is influenced by both activity dependent and independent factors Much of original structure is dictated by activity dependent factors Refinement comes from activity This is a result of LTP-like mechanism But in general, it’s hard to say which causes which feature…

Activity Independent Experiments Eye rotation in newt Rotation of the eye of an adult newt will cause the newt to see the world upside-down because in the adult brain, the retino-tectal connections don’t rewire, little plasticity. Previous projections from a part of the retina now project to the “wrong” part of the tectum. Retinal ablation Ablating half of the retina will cause missing connections in half of the tectum. The persisting retinal half will not rewire to take up the whole tectum. Stripe Assay Neurons from temporal retina will only grown onto membrane stripes from the anterior , and nasal retinal neurons will project through both (as it has to to get to the posterior tectum!)

Activity Dependent Experiments Rewiring of ferret cortex Rewiring of retinal projections to the MGN (after deafening the ferret) will cause A1 to have V1’s features like orientation pinwheels and long horizontal connection. Formation of eye specific stripes They don’t form if APV is perfused!

Mechanisms Ephrins and Eph Receptors Chemical gradient that guides neuronal projections from (e.g. retina to tectum) specific regions of one neural area to another specific region Axonal Segregation / Map Refinement Synapses that fire together wire together, so synapses become refined Accomplished via LTP (requires NMDAR activity) Synapse Maturation NMDA only synapses become unsilenced as a result of LTP (insertion of AMPARs) – change in NMDAR/AMPAR ratio Depolarizing GABA also aids in unsilencing Gene Expression e.g. cpg15 is induced by neural activity and regulates synaptic maturation GABA is depolarizing in immature synapses because of high levels of Cl- intracellularly in immature neurons (negative charges flowing out would cause depolarization).