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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. - PowerPoint PPT Presentation
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K+ Channel
Sukhee ChoGreg Richard
K+ Channels
• Found everywhere
• Contribute to resting potential (neurons)
• Major roles in cardiac tissue
• Involved in hormone secretion
ClosedOpen
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 A- type 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
http://en.wikipedia.org/wiki/Beta_amyloid
Alzheimer's diseased brain
Aβ1-40
Aβ1-42
Aβ1-40
Aβ1-42
Amyloid precursor protein
Controlling neurotransmitter releaseFast after-hyperpolarizationSpike frequency adaptation
VSD - voltage sensing domainPGD - pore-gating domainRCK - regulator of K conductance
Lee et al., Trends Neurosci. 2010 Sep;33(9):415-23. Review.
BK channelLarge conductance Ca2+-activated K+ channels, Maxi-K, BK or Bkca, Kca1.1
Figure 1. Intracellular infusion of Aβ1-42 broadens spike width and augmemted Ca2+ influx in rat neocortical pyramidal neurons.
Aβ1-40
Aβ1-42
Fura-2
100-250 pA500 ms
Figure 3. Intracellular Aβ1-42 enlarges spike width by suppressing BK channels, thereby increasing spike-induced Ca2+ entry.
Charybdotoxin - Ca2+-activated K+ channel blocker4-AP(4-Aminopyridine) – A-type potassium channel blocker
Figure 5. ECS blocked Aβ1-42-mediated suppression of BK channels in rat neocortical neurons.
Isopimaric acid
Electroconvulsive shock
Figure 7. Blocking effects of ECS on Aβ1-42 was absent in H1aKO mice.
Figure 8. Spike broadening in 3xTG neurons.
JuvenileJuvenile
4 months of age
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?
Trek2b
N-terminus
Trek2-1p
C-terminus
Trek2 Variants
Trek2
Immunoblotting
Myc-tag : N-EQKLISEEDL-C (1202 Da)
Whole-cell Currents(Voltage-step)
-100mV
+60mV
20mV
Reversal Potential (Erev)(Voltage-ramp)
-100mV
+60mV
1 s
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.
How does nature accomplish high conduction rates and high selectivity at the same time?
Sculpture by Julian Voss-Andreae
Roderick MacKinnon - 2003 Nobel Prize in Chemistry
Visualize a K+ channel and its selectivity filter
The signature sequence of the potassium channel
Yellow : carbon, Red : oxygen
Carbonyl oxygens attract K+ ions
Yellow : carbon, Blue : nitrogen, Red : oxygen
Electrostatic repulsion favors high conduction rates
Paper #3
http://radiographics.rsna.org
The renin-angiotensin-aldosterone system regulating blood pressure
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 70Most of them are classified as idiopathic and a small number have mutationsResulting in hypertension and hypokalemia (low plasma K+ level)Surgical procedure can relieve symptoms
Hereditary hypertension
Mendelian form of primary aldosteronismBilateral 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 channelKir3.4, GIRK4
Subclasses: ROMK, GPCR, ATP-sensitive2 TMD, 1 PCurrent 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.