lecture 6 - Electrophysiology Flashcards
What is electrophysiology?
Electrophysiological techniques are based around measuring this potential difference
membrane potentail
The membrane potential is the voltage across the neuronal membrane at any moment, represented by the symbol Vm.
The resting potential of a typical neuron is between -55 mV and -90 mV.
This is primarily driven by the permeability of potassium (K+) ions across the membrane
Why does membrane permeability to K+ ions lead to a negative membrane potential?
Membrane potential is dependent on
leak K+ channels
Membrane potential is dependent on leak K+ channels
The equilibrium potential
Also known as the ‘reversal potential’ or the ‘Nernst potential’
This is the membrane potential where the chemical and electrical gradients of an ion across a biological membrane are in equilibrium
At this membrane potential there is no net flow of ions across the membrane
Written as Eion (for example EK for the potassium equilibrium potential)
We can calculate the equilibrium potential of an ion across a membrane
The Nernst equation: simplified
Ionic equilibrium potentials
Each ion has a different equilibrium potential (i.e. the membrane potential at which there is no net flow of ions)
This is determined by:
The intracellular concentration
The extracellular concentration
The valance of the ion
Ion concentrations on either side of a neuronal membrane:
Calculate the equilibrium potential for the following ions
So how do I calculate the resting potential of a neuron?
You need various bits of information
Concentration of ions inside & outside the cell
The relative permeability of these ions
Plug them into the Goldman-Hodgkin-Katz (GHK) equation (modified from Nernst equation)
Worked example
How is the resting membrane potential maintained?
The sodium-potassium pump: Na-K ATPase pump
This ion pump is a membrane-associated protein that transports ions across the membrane against their concentration gradients at the expense of metabolic energy.
Sodium-potassium pump expends as much as 70% of the total amount of ATP utilized by the brain.
What happens to VM if we open a Na+ channel (eg by applying an channel agonist)?
What happens to VM if we open a K+ channel (eg by applying an channel agonist)?
What happens to VM if we open a Cl - channel (eg by applying an channel agonist)?
ECl : a moveable feast
ECl is usually close to the resting membrane potential
In adult neurons, usually ECl < Vrest, so activating Cl- channels usually hyperpolarises the membrane
In juvenile neurons, ECl > Vrest so activating Cl- channels depolarises the membrane
Why…?
Types of electrophysiology
Intracellular recordings of membrane potential
What preparation?It depends on the question you’re asking
Dissociated neuronal cultures
Pros: cells easily accessible for intracellular recording
Cons: no anatomical correlate; cells not in physiological environment; can only study early developmental stages
Acute brain slices
Pros: local circuits intact; can study any developmental stage; anatomically relevant
Cons: long range inputs/outputs severed; not physiological environment
Whole animal
Pros: all circuits intact; can correlate activity with behaviour
Cons: technically very challenging, especially for intracellular recordings
We use electrophysiology to measure the activity of ion channels
Ion channel subtype classification:
Pharmacology
Activation mechanism
Genetics
Carrier ion(s)
Na+
K+
Cl-
Ca2+
[There are others but these are most important]
NB – we can measure the effects of other types of receptors with ephys (e.g. G-protein coupled receptors), but only via their effects on ion channels
How do we study synapses using electrophysiology?
Usually record the activity in the postsynaptic neurone
Extracellular recording
Recap
Intracellular recordings: EPSP. Based on ion channels activities
Extracellular Recordings: fEPSP. Based on the stream of ions going inside the celles.
EPSP = synaptic activity of one single cell
fEPSP = synaptic activity of many synapses in many cells
How do we study the properties of ion channels using electrophysiology?
The problem
We want to study the voltage-dependence of a voltage gated ion channel (e.g. a Na+ channel underlying an action potential or a glutamate receptor underlying an EPSP).
For example what membrane potential does it open at?
With a ‘current clamp’ recording, by definition the membrane potential is constantly changing.
The solution
We fix the voltage at particular membrane potentials and study the underlying current – This is ‘voltage clamp’
CURRENT clamp: measures the membrane potential
VOLTAGE clamp: records the current
Current clamp’ – measuring the membrane potential (voltage)
Electrophysiological amplifiers are capable of stimulating the cell by injecting current.
Therefore, intracellular recordings of voltage are called ‘current clamp’
Imagine you are trying to maintain your bank balance at exactly £0…
Voltage clamp experiment studying synaptic currents
Summary
Intracellular electrophysiology used to study electrical activity in single cells
Extracellular electrophysiology used to study electrical activity from groups of cells in one anatomical location
Current clamp recording can be used to monitor the effects of manipulations (eg drugs) on membrane potentials
Voltage clamp can be used to study the underlying biophysical properties of channels in the membrane and the effects of manipulations (eg drugs) on these
