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Action Potential 1

Physiology and Pharmacology

QuestionAnswer
What is the Electrical Analogue of the cell membrane Batteries = concentration gradients of ions Conductance = ion channels in the membrane allowing the movement of ions. Movement of Na+ generates a positive current Movement of K+ generates a negative current
Basic process of an action potential Triggered by membrane depolarisation above a stimulus threshold ( when Na influx is greater that K efflux). The amplitude is all or nothing with a peak potential of up to 40 mV with a variable duration. Followed by hyperpolarisation refractory period)
Name and describe the two types of refractory period Absolute refractory period - period when it is impossible to generate an action potential no matter how strong the stimulus Relative refractory period - weak action potentials can be generated but a larger stimulus is needed
Mechanism of an action potential Caused by increased conductance of Na+ as more channels open and a large influx of Na+ causes depolarisation This opens voltage gated K+ channels as the Na+ channels close and there is a large efflux of K+ to repolarise the membrane.
Evidence for the role of Na+ and K+ AP fails in Na free solution with reduction of Na producing reduction in AP Radiotracers show Na entry and K efflux Genes and proteins for the channels allow understanding of how they work. This is supported by x-ray crystallography
How a voltage clamp works Voltage is set via a clamp, with actual cell voltage measured. Current is injected into the cell proportional to the difference in set and actual voltage. This encouraged depolarisation so ionic currents can be measured
Hodgkin and Huxley - Voltage clamp depolarisation of a squid axon Upon depolarisation there was a inward flow of positive charge into the axon, which is followed by an outwards current of positive charge. By removing Na+ from the solution they isolated the K+ effects (the outwards current as no inward current seen).
Hille - Isolating K+ and Na+ currents Using a tetrodoxin inhibitor for Na channels - no inwards current seen, only the outwards K current is present Using a Tetraethylammonium inhibitor for K voltage gated channels no outwards currents seen, only the inwards Na current is present
Hille's findings on strength of response K+ - as depolarisation increased a larger response was seen Na+ - as depolarisation increased response rapidly increased as all Na channels were activated. Once all were activated further depolarisation decreased response as channels began to close
Positive feedback in action potentials Small depolarisation of the membrane opens Na voltage gated channels. This depolarises the membrane further, opening more Na voltage gated channels in a positive feedback loop
Negative feedback in action potentials Depolarisation activates K+ voltage gated channels. K+ moves out the cell beginning to repolarise it, causing some of the open channels to close. This closure of channels in a negative feedback loop is slow, which is why membranes may hyperpolarise
Inactivation of voltage gated sodium channels Following activation Na channels inactivate, Na cannot pass through these channels and they cannot reopen until the membrane is fully repolarised. This does not occur instantaneously, which leads to the refractory period
Hille - Time dependence of recovery from inactivation Varied time delays between action potentials. With a short delay no Na current was observed in the second depolarisation, whilst with longer delays Na currents began to be observed again. Overall conclusion - full recovery from activation occurs at 10 ms
Process of an action potential Rapid voltage dependant activation of Na channels allows an influx of Na+. This depolarises the membrane. The Na channels then inactivate and K voltage gated channels open slowly (delayed rectifiers). K moves out the cell to repolarise it
What is threshold potential Voltage at which AP is triggered, occurs when inwards Na current is greater than outwards K leak currents
What is All or nothing response Once threshold is reached activation of Na channels causes depolarisation with activates a positive feedback loop
What causes hyperpolarisation Voltage activated K+ channels are slow to close on repolarisation, so membrane potential is initially more negative immediately after repolarisation
Studying single channels - Patch clamp Press a glass pipette to a membrane and suck to create an electrically tight seal. Can be studied like this or by pulling the electrode away from the cell to create an inside out patch.
Hille - findings from Patch clamp Na channel-current always the same magnitude but when a channel opens is a random process. They generate an inward current with rapid action K channel-active through the whole repolarisation but are not open the whole time. Generate an outward current
Voltage gated Na+ channels A transmembrane spanning protein with a pore through the centre to allow ions through. Narrow space of the pore is a selectivity filter, forming bonds with the ion to dehydrate it. There is a voltage sensing region coupled to an activation gate
Sequence of Na+ channel gating At rest- Activation gate is closed Depolarisation - voltage sensor moves to open the activation gate Inactivated - Inactivation gate blocks to pour to close it
Structure of a Na+ channel alpha subunit There are 4 similar domains each with 6 alpha helices that span the membrane. S4 is the voltage sensing helix with charged AAs, whilst S5 and S6 form the centre of the channel. They are connected by a semi helical structure
Structure of a K+ channel Made of 4 individual subunits with loops between S5 and S6 which contain a specific sequence of amino acids to allow electrostatic interaction with ions. There is a crossing of different subunits at the inner helix to generate the gate
Paddle domains on Voltage gated channels These contain the voltage sensing region. Their location is determined by the membrane potential. They are either attracted to or repelled by the membrane potential, changing where the rest. Their movement opens and closes the gate
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