Week 1 - Channels and Transporters

Question 1

what is the voltage patch-clamp method? what does it tell us about channels and gating?

Answer 1

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

Question 2

macroscopic and microscopic methods

Answer 2

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)

Question 3

patch clamp measurements of ionic currents through single Na+ channels, both macroscopically and microscopically

Answer 3

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

Question 4

sustained response for K+ channels

Answer 4

on average, K+ channels tend to be an open state while the membrane is depolarized

Question 5

K+ compared to Na+ channels

Answer 5

K+ are in the opposite direction, with longer latency for activation and long duration of activation

Question 6

summary of patch clamp (microscopic) and voltage-clamp (macroscopic) Na+ and K+ channels

Answer 6

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+

Question 7

what do multiple potassium channel types do?

Answer 7

add diversity

1. most CNS neurons have multiple K+ channels with different characteristics
2. voltage dependence of activation (low-voltage VS high voltage activation)
3. rate of activation (how fast population reaches max conductance)
4. inactivation properties (creates diversity of spike waveforms and spike patterns for different cells)

Question 8

functional roles of fast after hyperpolarization

Answer 8

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

Question 9

functional roles of medium after hyperpolarization

Answer 9

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

Question 10

functional roles of slow after hyperpolarization

Answer 10

100-3000 ms; limits firing frequency by unknown channel

Question 11

channel timings in neurons

Answer 11

1. Na+ channels open
2. Na+ channels inactivate, Ka+ and Kdr+ channels open
3. Kbk+ channels open
4. Ca++ channels open
5. other known and unknown K+ channels open

Question 12

how many types of ions pass through voltage gated and ligand gated channels?

Answer 12

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

Question 13

how are neurons with diverse electrical properties created?

Answer 13

large numbers of ion channel genes

• 10+ for Na+ and Ca++ channels, 100+ for K+
• splicing variations produce different characteristics

Question 14

how are ionic channels organized?

Answer 14

based on sequence homology
-voltage-dependent ion channels differ in cellular expression and subcellular localization impacting their relative contribution to brain function

Question 15

what do Kv4.1 channels do?

Answer 15

play a positive role in tumorigenic human mammary cells

Question 16

difference between Kv1.4 and Kv2.1

Answer 16

1. 4 - axons; in terminal fields of medial perforant path in middle molecular layer of dentate gyrus, and mossy fiber axons and terminals
2. 1 - soma and proximal dendrites; most prominent in pryamidal cell CA-1 layer

Question 17

why are there so many genes encoding K+ channels?

Answer 17

so the channels can differ in:
-activation
-gating
-inactivation
and shape complex electrical responses
-influence duration of AP and resting membrane potential

Question 18

relationship between Kv2.1, Kv4.1, inward rectifier channels, and Ca++ activated K+ channels

Answer 18

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

Question 19

summary of ion channel properties

Answer 19

• 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

Question 20

what are channelopathies?

Answer 20

genetic diseases due to mutations in channel genes

Question 21

what are channelopathies from mutations in voltage-gated Ca++ channels?

Answer 21

• congenital stationary night blindness
• familial hemiplegic migrane
• episodic ataxia type 2

Question 22

what are channelopathies from mutations in Na+ channel?

Answer 22

generalized epilepsy with febrile seizures

Question 23

what are channelopathies from mutations in K+ channels?

Answer 23

benign familial neonatal convulsion

Question 24

what are ion channel target sites for toxins?

Answer 24

extracellular domains and pore regions

Question 25

what does tetrodotoxin do?

Answer 25

block Na+ channels

Question 26

what does saxitoxin (red tide)

Answer 26

blocks Na+ channels (homologue of TTX)

Question 27

what do alpha and beta toxins from scorpions do?

Answer 27

alpha - prolong duration of Na+ currents

beta - shift voltage activation of Na+ channels

Question 28

what does batrachotoxin (frogs) do?

Answer 28

inactivation of Na+ channels (used by South American Indians)

Question 29

what do dendrotoxin (wasps) and apamin (bees) do?

Answer 29

K+ channel blockers

Question 30

what do conotoxins (cone snails) do?

Answer 30

block N-type Ca++ channels

Question 31

what do agatoxin (spiders) do?

Answer 31

block P/Q-type Ca++ channels

Question 32

what are active ion transporters and what do they do?

Answer 32

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

Question 33

where do ATPase pumps, ion exchangers, and co-transporters get energy from?

Answer 33

ATPase pumps get energy from ATP hydrolysis

ion exchangers and co-transporters get energy from electrochemical gradient on other ions

Question 34

what does electrogenic mean?

Answer 34

don’t pass the same number of ions on each side, and don’t contribute significantly to currents

Question 35

what are mechanoselective ion channels?

Answer 35

sense deformation in the membrane

• touch
• hearing
• osmoregulation
• neuromuscular stretch

Question 36

what are heat sensitive ion channels

Answer 36

• pain
• temperature
• inflammatory response