1. From Hodgkin (1957)

2. Ca2+-dependent action potentials

3. Ca2+ action potential properties
(Hagiwara & Byerly, 1978)
Overshoot or rate of rise varies with Cao (29 mV per 10-fold)
AP disappears when Mg replaces Ca
Overshoot and rate of rise don’t change when Nao removed
AP persists when Ca replaced with Sr or Ba
AP blocked by metal cations:
Co2+, La3+, Mn2+, Cd2+, Ni2+

4. Ca2+ currents are more difficult to study than Na+ currents
Current separation not simple: Ca2+ current overlaps with K+ current
K+ current often depends on ICa (Ca2+-activated K+ current)
No defined Ca2+ equilibrium potential: ECa not measurable
can’t use I = g(V-ECa)
Inactivation is complex: depends on current as well as voltage
Voltage-dependent facilitation: repetitive voltage steps can increase current
Dependence on phosphorylation: washout during internal perfusion (NT changes with time)

5. Na+ current
Ca2+ current
[Ca2+]o/[Ca2+]i = ~10,000
[Na+]o/[Na+]i = ~15
Erev = ~+50 mV
Erev not measurable
conductance is f(V, Ca2+)
linear conductance
Ion saturation and block
Ion fluxes are independent

6. Ca2+ conductance is not linear
Rectification:
Ca2+ equilibrium potential
not measurable
conductance large at negative Vm
conductance small at positive Vm
[Ca2+]inexp(2VF/RT)-[Ca2+]o
iCa(V) = PCa 4F2 V RT
exp(2VF/RT) - 1

7. How to get high selectivity and high flux rate
Problem:
How to get high selectivity and high flux rate
Na+
Na+ ionic radius 1.14 Å
Ca2+
Ca2+ ionic radius 1.16 Å
[Na]o >> [Ca2+]o
iCa = 106 ions/second
out in

8. Rate of rise of Ca action potential
Ion saturation indicates Ca2+ binds to the channel
Ba2+
Rate of rise of
Ca action potential
Ca2+
KBa ~ 20 mM
KCa ~ 5 mM
Concentration (mM)
Saturation
ICa ~ 1
1 + [Ca2+] KD
Ca binds to the channel with mM KD
How to reconcile low affinity with high selectivity

9. Ca2+ channels let Na+ ions through in the absence of Ca2+
(Almers et al, J. Physiol. 353:565, 1984)
I-monovalent
Cao = 60 nM
30 µM Cao
Cao = 30 µM
Imonovalent
blocked by
µM Cao
60 nM Cao
High-affinity Ca2+ binding site in channel

10. Outward current through Ca2+channels carried by monovalent cations
(Lee & Tsien, J Physiol. 354:253, 1984)
Experiment
Out: Ba
In: Cs
Whole-cell
large Iout
Cs+
Ba2+

11. Problem:
Low affinity site (millimolar) from saturation of ICa
High affinity site (micromolar) from block of monovalent INa+ by Ca2+o

12. Two binding site model for Ca2+ channel permeation
-log[Ca]

13. Ca2+ currents produce voltage-dependent Ca2+ accumulation
snail
neuron
snail
neuron
snail
neuron
salamander
photoreceptor
squid
synapse

14. Inactivation depends on Ca2+ current

15. Two-pulse voltage clamp analysis of current-dependent inactivation
Ca2+ current
during prepulse
(normalized to max)
Ca2+ current during
second test pulse
(normalized to max)
Ca2+ current during test
pulse reduced maximally
when prepulse current
evokes max Ica
(Ba2+ current not affected)

16. Ion channel diversity: why?
Resting potential
Complex electrical activity:
Pacemaker potentials
Burst generation
Spike adaptation
Resonance
Amplification of synaptic responses
Coupling electrical activity to neuronal function:
Ca2+ entry: neurotransmitter release,
growth cone motility, gene expression
Calcium waves and oscillations
Development
Impedance matching

18. Molecular diversity of voltage-dependent ion channels
Name
Genes
Physiologically defined currents K+ Kv 27
Neuronal delayed rectifier; A-current
Eag 8
Cardiac fast ‘delayed rectifier’
KCNQ 5
M-current; cardiac slow ‘delayed rectifier’ SK 4
Low conductance Ca2+-dependent
Slo 3
High conductance Ca2+- and voltage-dependent
Kir 16
Inward rectifiers, G protein-activated 2P 15
Leakage (resting potential)
Na+
TTX-s 6
Neurons, adult skeletal muscle, glia
TTX-r
Cardiac muscle, some sensory neurons
Ca2+
S, C, D, F
High voltage-activated (L-type)
A, B, E
High voltage-activated (N, P/Q, R-types)
G, H, I
Low voltage-activated (T-type)
Cation
Pacemaker
Cardiac muscle, some neurons
CNG
Cyclic nucleotide-gated
Trp 17
Capacitative Ca2+ entry

19. Na+ channel diversity
TTX-resist.
INa fast: H-H kinetics
INa persistent: incomplete inactivation
INa resurgent: passes through open state during recovery from inactivation
(NaV1.6)

20. Na+ currents in DRG neurons
Large afferent
Fast
TTX-sens.
Nav1.6
Small sensory
Slow
TTX-resist.
NaV1.7,8,9
Incomplete inactivation:
Vm = -30
m = ~.3
h = ~.3
Small sensory
Both
contributes to
hyperexcitability

21. Persistent Na+ currents amplify sub-threshold synaptic responses
cortical & hippocampal pyramidal cells
Persistent INa amplifies synaptic potential
hyperpolarize
cell

22. Resurgent Na+ currents in cerebellar Purkinje neurons
(Raman & Bean, 1997)
fast INa

23. Resurgent Na+ currents: burst generation
R  O  I
Resurgent I  O recovery

24. Ca2+ channel diversity Nifedipine, diltiazem, verapamil
Dihydropyridine-
sensitive
Nifedipine, diltiazem,
verapamil
Dihydropyridine-
insensitive
agatoxin IVA, conotoxin MVIIC
w-conotoxin GVIA
‘resistant’
P/Q, N, and R: presynaptic localization, transmitter release
L-type: postsynaptic localization, cell bodies and dendrites
gene expression, cerebellar LTD, plateau potentials

25. High- and low threshold Ca2+ currents
Weak depol
Strong depol
Large, sustained
Inward Ba current
LVA - T-type
HVA - L, N, P, R

26. Burst firing in thalamocortical relay neurons: T-type Ca2+ channels (CaV3.1)

27. Conus striatus - a fish eater

28. Pharmacological components of neuronal high-threshold Ca2+ currents
Acutely isolated cortical & striatal neurons
(Mermelstein et al, J. Neurosci. 19:7268, 1999) L P N Q

29. K+ channel diversity charybdotoxin apamin dendrotoxin
tetraethylammonium ion
Ba2+

30. Different K+ currents contribute to action potential complexity

31. Rapidly inactivating K+ current (IA)
(Connors & Stevens, 1971)

32. IA controls bursting in snail neurons
(Connors & Stevens, 1971)
IA contributes to maintained hyperpolarization
Repolarization removes IA inactivation

33. Ca2+-activated K+ current (IK-Ca)
(Meech, 1970s) mV IK
- Cao mV
+150
Slope = 29 mV/10-fold ∆Cao
IK-+Ca- IK-Ca Er
Cao
5 Ca
10 Ca
20 Ca

34. Ca2+-activated K+ currents contribute to the after-hyperpolarization

35. Ca2+accumulation and Ca2+-activated K+ current

36. Resonance in hair cells
(Art & Fettiplace, J Physiol 385: , 1987)

37. Inwardly rectifying K+ currents
Hyperpolarization-activated
Gating depends on V-EK
Rectification due to block by internal Mg2+
Functions: Sets resting potential
Augments depolarization & repolarization
Protects cells against damage-induced depolarization

38. Pacemaker channels
HCN hyperpolarization-activated
Cation-permeable