Week 1 - Channels and Transporters Flashcards
what is the voltage patch-clamp method? what does it tell us about channels and gating?
detects currents flowing through single membrane channels due to depolarization
- separate Na+ and K+ channels
- voltage sensors in channels
- high conductance of channels to specific ions
- while channels open and close in all-or-none fashion with a fast switch between open/closed states in stochastic manner, gatings transition between open and closed states and involve temporary conformational change in channel’s structure
macroscopic and microscopic methods
macroscopic (voltage-clamp) currents: due to flow through many channels (whole-cell recording; strong pulse of suction so cytoplasm is continues with pipette interior; the pipette is still attached to the membrane)
microscopic (patch clamp) currents: due to current flow through one channel (inside-out recording; exposed to air and cytoplasmic domain is accessible within the pipette)
patch clamp measurements of ionic currents through single Na+ channels, both macroscopically and microscopically
depolarization increases probability of a channel being open, and hyperpolarization decreases it
- K+ channels were blocked via TEA
- comparing the macro and micro, it showed that the sum of many trials microscopically was correlated with the macroscopic trial
sustained response for K+ channels
on average, K+ channels tend to be an open state while the membrane is depolarized
K+ compared to Na+ channels
K+ are in the opposite direction, with longer latency for activation and long duration of activation
summary of patch clamp (microscopic) and voltage-clamp (macroscopic) Na+ and K+ channels
Na+ opening is voltage-dependent, near beginning of depolarization pulse
- they inactivate, current reverses at E Na
- blocked by TTX
K+ opening is voltage-dependent, and open later
- many don’t inactivate, but merely close
- blocked by TEA or Cs+
what do multiple potassium channel types do?
add diversity
- most CNS neurons have multiple K+ channels with different characteristics
- voltage dependence of activation (low-voltage VS high voltage activation)
- rate of activation (how fast population reaches max conductance)
- inactivation properties (creates diversity of spike waveforms and spike patterns for different cells)
functional roles of fast after hyperpolarization
2-5 ms; shortens AP by quickly repolarizing membrane
- only affects early spike frequency at very high frequencies
- big K+ channels activation by Ca++ and depolarization, then rapid inactivation
functional roles of medium after hyperpolarization
10-100 ms; controls early interspike interval
- contributes to early spike-frequency adaptation by slowly activating Ca++ entry
- controls late spike-frequency adaptation
- intermediate and small K+ channels are non-inactivating
functional roles of slow after hyperpolarization
100-3000 ms; limits firing frequency by unknown channel
channel timings in neurons
- Na+ channels open
- Na+ channels inactivate, Ka+ and Kdr+ channels open
- Kbk+ channels open
- Ca++ channels open
- other known and unknown K+ channels open
how many types of ions pass through voltage gated and ligand gated channels?
voltage gated channels allow only a single type of ion to pass through the channel, though there are exceptions
ligand gated ion channels usually allow 2+ types of ions to pass through the channel
how are neurons with diverse electrical properties created?
large numbers of ion channel genes
- 10+ for Na+ and Ca++ channels, 100+ for K+
- splicing variations produce different characteristics
how are ionic channels organized?
based on sequence homology
-voltage-dependent ion channels differ in cellular expression and subcellular localization impacting their relative contribution to brain function
what do Kv4.1 channels do?
play a positive role in tumorigenic human mammary cells
difference between Kv1.4 and Kv2.1
- 4 - axons; in terminal fields of medial perforant path in middle molecular layer of dentate gyrus, and mossy fiber axons and terminals
- 1 - soma and proximal dendrites; most prominent in pryamidal cell CA-1 layer
why are there so many genes encoding K+ channels?
so the channels can differ in: -activation -gating -inactivation and shape complex electrical responses -influence duration of AP and resting membrane potential
relationship between Kv2.1, Kv4.1, inward rectifier channels, and Ca++ activated K+ channels
Kv2.1 - little inactivation, and are related to channels involving in repolarization
Kv4.1 - inactivate rapidly to depolarization
IRC - allow more current flow during hyperpolarization than during depolarization
CAKC - open in response to increased intracellular Ca++ and sometimes to membrane depolarization
summary of ion channel properties
- encoded by large and diverse families of homologous genes
- differ widely in cellular expression and subcellular localization
- different voltage-gated channels differ in functional properties (activation, inactivation, gating)
- contribute to complex (rich) electrical responses
- their diversity is key to developing new therapeutics for CNS disorders
what are channelopathies?
genetic diseases due to mutations in channel genes
what are channelopathies from mutations in voltage-gated Ca++ channels?
- congenital stationary night blindness
- familial hemiplegic migrane
- episodic ataxia type 2
what are channelopathies from mutations in Na+ channel?
generalized epilepsy with febrile seizures
what are channelopathies from mutations in K+ channels?
benign familial neonatal convulsion
what are ion channel target sites for toxins?
extracellular domains and pore regions
what does tetrodotoxin do?
block Na+ channels
what does saxitoxin (red tide)
blocks Na+ channels (homologue of TTX)
what do alpha and beta toxins from scorpions do?
alpha - prolong duration of Na+ currents
beta - shift voltage activation of Na+ channels
what does batrachotoxin (frogs) do?
inactivation of Na+ channels (used by South American Indians)
what do dendrotoxin (wasps) and apamin (bees) do?
K+ channel blockers
what do conotoxins (cone snails) do?
block N-type Ca++ channels
what do agatoxin (spiders) do?
block P/Q-type Ca++ channels
what are active ion transporters and what do they do?
membrane PRO that create and maintain ion gradients
- form complex w/ ion they transport
- binding and unbinding is slow (in milliseconds)
- ion translocation is slower in transporters than in channels
where do ATPase pumps, ion exchangers, and co-transporters get energy from?
ATPase pumps get energy from ATP hydrolysis
ion exchangers and co-transporters get energy from electrochemical gradient on other ions
what does electrogenic mean?
don’t pass the same number of ions on each side, and don’t contribute significantly to currents
what are mechanoselective ion channels?
sense deformation in the membrane
- touch
- hearing
- osmoregulation
- neuromuscular stretch
what are heat sensitive ion channels
- pain
- temperature
- inflammatory response
