Potassium Ion Channel - FSU Program in Neuroscience

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K
Channel
Sukhee Cho
Greg Richard
K+ Channels
• Found everywhere
• Contribute to resting potential (neurons)
• Major roles in cardiac tissue
• Involved in hormone secretion
Open
Closed
Slow to close
Inactivated
K+ Channel Anatomy
Senyon Choe (2002)
Gating
Bezanilla 2004
Classes
• Inwardly Rectifying
– ROMK, GIRK, ATP-sensitive
• Tandem Pore Domain
– TWIK, TREK, TASK, TALK, THIK, TRESK
• Voltage-Gated
– hERG, KvLQT1
• Calcium Activated
– BK, IK, SK
Inwardly Rectifying (Kir, IRK)
• Subclasses: ROMK, GIRK, ATP-sensitive
• 2 TMD, 1 P
• Current flow into cell (“inward”)
• Differ from delayed rectifier or Atype channels (outward current)
Tandem Pore Domain (K2P)
• Subclasses: TWIK, TREK, TASK, TALK, THIK, TRESK
• 4 TMD, 2 P (two 2 TMD, 1 P)
• “Leak channels” – contribute to resting potential
• Activated by mechanical stretch, pH, temperature
Voltage-Gated (Kv)
• Subclasses: hERG, KvLQT1
• 6 TMD, 1 P
• Sensitive to voltage changes
– S4 domain
• Return to resting state
– Repolarization
– Limits AP frequency (RRP)
Calcium Activated (KCa1 )
• Subclasses: BK, IK, SK
• 6 TMD, 1 P
• Activated by intracellular Ca2+
• Some activated by intracellular Na+ & Cl• N-terminus extracellularly (Unlike Kv)
Paper #1
Amyloid β Hypothesis in Alzheimer’s disease
Aβ1-40
Aβ1-42
Alzheimer's diseased brain
Amyloid precursor protein
http://en.wikipedia.org/wiki/Beta_amyloid
BK channel
Large conductance Ca2+-activated K+ channels, Maxi-K, BK or Bkca, Kca1.1
VSD - voltage sensing
domain
PGD - pore-gating
domain
RCK - regulator of K
conductance
Controlling neurotransmitter release
Fast after-hyperpolarization
Spike frequency adaptation
Lee et al., Trends Neurosci. 2010 Sep;33(9):415-23. Review.
Aβ1-40
Aβ1-42
Fura-2
500 ms
100-250 pA
Figure 1. Intracellular infusion of Aβ1-42 broadens spike width and augmemted
Ca2+ influx in rat neocortical pyramidal neurons.
Charybdotoxin - Ca2+-activated K+ channel blocker
4-AP(4-Aminopyridine) – A-type potassium channel blocker
Figure 3. Intracellular Aβ1-42 enlarges spike width by suppressing BK channels,
thereby increasing spike-induced Ca2+ entry.
Isopimaric acid
Electroconvulsive shock
Figure 5. ECS blocked Aβ1-42-mediated suppression of BK channels in rat
neocortical neurons.
Figure 7. Blocking effects of ECS on Aβ1-42 was absent in H1aKO mice.
4 months of age
Juvenile
Juvenile
Figure 8. Spike broadening in 3xTG neurons.
Figure 9. Recovery of single BK current by ECS in 3xTG mice.
Conclusions
Intracellular Aβ1-42 broadens spike width in neocortical
pyramidal neurons by downregulation of BK channel
activities.
ECS counteracts Aβ1-42 induced BK channel inhibition by
expression of Homer 1a
Paper #2
Trek Channels
• Two-pore domain K+ channels (K2P)
– 4 TMD, 2 pore
• Subfamilies:
– Trek1 (Kcnk2)
– Trek2 (Kcnk10)
• Underlie “leak” and background K+ conductances
• Sensitive to membrane stretch, temperature, & pH
• Inhibited by PKC & PKA
Trek2
• Trek2b
– Differs from Trek2a & Trek2c at N-terminus
• Trek2-1p
– C-terminal truncation (2 TMD & 1 pore)
Does alternative splicing of Trek2 contribute to functional
diversity of channel as seen with Trek1?
Trek2 Variants
Trek2
Trek2-1p
C-terminus
Trek2b
N-terminus
Immunoblotting
Myc-tag : N-EQKLISEEDL-C (1202 Da)
Whole-cell Currents
(Voltage-step)
+60mV
20mV
-100mV
Reversal Potential (Erev)
(Voltage-ramp)
+60mV
1s
-100mV
Non-selective channel
Whole-cell Currents
Surface Trek2 Expression
Total Protein
Surface Protein
Conclusions
• Trek2b exhibited larger currents than Trek2b & 2c; > # of Trek2b
channels on membrane surface.
• As [K+]o , Erev ; overexpression of K+-selective channels
• Trek2-1p may require additional assembly to form functional
channels.
• N-terminal variation can influence current amplitude and
surface level of Trek2 channels, as seen in Trek2b.
Sculpture by Julian Voss-Andreae
How does nature accomplish high conduction
rates and high selectivity at the same time?
Visualize a K+ channel and its selectivity filter
Roderick MacKinnon 2003 Nobel Prize in
Chemistry
The signature sequence of the potassium channel
Carbonyl oxygens attract K+ ions
Yellow : carbon, Red : oxygen
Electrostatic repulsion favors high conduction rates
Yellow : carbon, Blue : nitrogen, Red : oxygen
Paper #3
The renin-angiotensin-aldosterone system
regulating blood pressure
http://radiographics.rsna.org
The angiotensin-renin-aldosterone system
regulating blood pressure
Adrenal glomerulosa cells
in the zonaglomerulosa
Choi et al., Science
Aldosterone-producing adenomas (Aka Conn’s syndrome)
One of the most common types of the primary aldosteronism (the overproduction of
aldosterone)
Conn’s sydrome is caused by a discrete benign tumor of the adrenal gland (APA)
Diagnosed between ages 30 and 70
Most of them are classified as idiopathic and a small number have mutations
Resulting in hypertension and hypokalemia (low plasma K+ level)
Surgical procedure can relieve symptoms
Hereditary hypertension
Mendelian form of primary aldosteronism
Bilateral adrenal hyperplasia (increase in number of cells/proliferation of cells)
Bilateral adrenalectomy in childhood
Protein-changing somatic mutations in aldosterone-producing adenomas
Mutations in KCNJ5 in aldosterone-producing adenoma and inherited aldosteronism
The probability of seeing either of two somatic mutations recur by chance in 6 of 20 other tumors is <10-30
H.s., Homo sapiens Human
M.m., Mus musculus Rodent
G.g., Gallus gallus Chicken
X.t., Xenopus tropicalis Frog
D.r., Danio rerio Zebrafish
C.I., Ciona intestinalis Sea squirt
KCNJ5 channel
Kir3.4, GIRK4
Subclasses: ROMK, GPCR, ATP-sensitive
2 TMD, 1 P
Current flow into cell (“inward”)
Differ from delayed rectifier or A-type channels (outward current)
Magnesium ions, that plug the channel pore at positive potentials, resulting in a
decrease in outward currents.
A voltage-dependent block by external Cs+ and Ba2+
Location of human mutations in KCNJ5 mapped onto the crystal
structure of chicken K+ channel KCNJ12
KCNJ5 mutations result in loss of channel selectivity and membrane depolarization
KCNJ5 mutations result in loss of channel selectivity and membrane depolarization
Membrane depolarization by either elevation of extracellular K+ or
closure of K+ channels by angiotesin II activates voltage-gated Ca2+
channels, increasing intraceullular Ca2+ level.
Channel containing KCNJ5 wit G151R, T158A, or L168R mutations
conduct Na+, resulting in Na+ entry, chronic depolarization,
constitutive aldosterone production, and cell proliferation.
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