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epithelial > L8 - K+ channels > Flashcards

L8 - K+ channels Flashcards

1
Q

what is the main function of K+ in cells?

A
  • drive the cell to a negative resting potential (drive Vm to Ek)
  • high IC K+ and low EC K+
  • when K+ channels activated, membrane potential becomes more negative - this creates a driving force for Cl- secretion
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2
Q

give the 3 main families of K+ channels in epithelia

A
  • voltage-gated K+ channels (Kv)
  • inwardly rectifiying K+ channels (Kir)
  • 2 pore domain K+ channels
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3
Q

give some features of voltage-gated K+ channels

A
  • Po changes with membrane potential
  • 6TMD and 1 pore (between TMD5-6)per subunit
  • 4 subunits make up a functional channel
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4
Q

Give some features of a inwardly rectifying potassium channel

A
  • greater tendency for K+ uptake
  • 2TMD and 1 pore per subunit
  • 4 subunits make up a functional channel
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5
Q

Give some features of a 2 pore domain potassium channel

A
  • 4 TMD per subunit
  • 2 pores per subunit
  • 2 subunits make up 1 channel
  • are CONSTITUTIVELY ACTIVE - help set resting potential
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6
Q

How many pore sequences to each different types of K+ channels have?

A
  • each channel has 4 pore sequences
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7
Q

what is the hypothesis to do with K+ channels and Cl- secretion?

A

K+ maintains Cl- secretions, as open K+ channels cause K+ uptake into the cell - causes hyperpolarisation - creates driving force for Cl- secretion via CFTR

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8
Q

what does chromanol 293B inhibit?

A

inhibits small number of K+ channels, e.g. KCNQ1 gene

- KvLQT1 channel

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9
Q

Is there any difference between the levels of KVLQT1 in normal and CF tissue? how was this shown?

A

No, there was no difference between mRNA levels expressed in the upper respiratory tract between normal and CF tissues
- shown doing RT-PCR of RNA in HBE cells line

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10
Q

what did experiments using a ussing chamber on nasal epithelial cells show about Cl- secretion and K+?

A
  • When levels of chromanol 293B rise - there is a change is Vte - this shifts more towards 0 which shows that there is a decrease in Cl- secretion
  • cannot be due to changes in K+ as ussing chamber doesn’t measure recycling of K+ - only measures net current
  • cannot be due to ENaC- as amiloride is added to avoid interference
  • size of CFTR currents reduce with increased 293B as there is a reduced driving force for cl- secretion
  • but is still some Cl- secretion as it is not all blocked by 293B - must be other channels driving Cl- secretions
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11
Q

What happens when Ba2+ is added in exp with ussing chamber measuring Cl- secretion in nasal epithelia?

A
  • when Ba2+ is added - this blocks most k+ channels - makes graph/current drop to 0 - must be other channels driving Cl- secretion which are inhibited by Ba2+ but not by chromanol 293B
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12
Q

How was the response to chromanol 293B effected in CF tissue compared to WT?

A
  • when IBMX/fsk to WT the response to chromanol 293B was larger than without IBMX.fsk - shows that more Cl- secretion is going on
  • In CF tissues - no change in Cl- secretion when 293B added as no CFTR channels are working anyway
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13
Q

How was the response to Ba2+ effected in CF tissue compared to WT?

A
  • In CF tissue there was a bigger change in Isc when Ba2+ was added compared to when 293 B is added
  • shows Cl- secretion must be driven by Ba2+ sensitive K+ channels, and that there must be another form of Cl- secretion apart from CFTR- as CFTR is non-functional in CF patients - but is still Cl-secretion occurring
  • also no change in Isc to IBMX/fsk - shows secretion is not due to CFTR
  • is no sig difference between CF and WT tissues - both have other mechanism of Cl secretion
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14
Q

what is the basolateral k+ channel that drives Cl-secretion through CFTR?

A
  • is KvLQT1 (Q1) with E3 - this is cAMP activated - can drive Cl- secretion through CFTR
  • is chromanol 293B sensitive
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15
Q

What is the other K+ channel on the basolateral membrane of epithelila cells which can drive Cl- secretion?

A

hSK4

  • is a Ca2+ activated K+ channel
  • is barium sensitive
  • is blocked by clotrimazole
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16
Q

what is another Cl- channel on the apical membrane that is responsible for the small amount of Cl- secretion in CFTR patients?

A

CaCC

  • Ca2+ activated Cl channel
  • activated by UTP -this increases IC Ca2+
  • purinoceptors activate UTP
17
Q

what Cl- channel is upregulated in CF and how was this shown?

A
  • CaCC is upregulated in CF patients
  • Showed as apical UTP increase in ISc was twice as much in CF tissue nasal as in WT tissue - higher Cl- secretion
  • when UTP added - is a hyperpolarising shift
  • this gives slight compensation for lack of CFTR
  • Is Po or number increased?
18
Q

what is the K+ channel that drives UTP dependent Cl- secretion? how was this shown?

A

hSK4
- when SK4 was inhibited by clotrimazole there was no/little UTP induced Cl- secretion
- HSK must be providing driving force for Cl- secretion via CaCC channels.
(when 293B added- no change - shows Cl- secretion is not due to Q1)

19
Q

what can inhibit CA2+ activated Cl- secretion ?

A

clotrimazole

20
Q

can Q1 channels switch?

A

propose that Q1 channels switch in CF and drive Cl- secretion through CaCC not CFTR

21
Q

is there controversy in effects Q1 has on Cl- secretion in CF?

A

Yes

  • some exp show that addition of chromanol 293B has no change on Isc - Q1 having no effect/ contribution to Cl- secretion in CF
  • other exp show there is a reduction in UTP induced current (Cl- secretion) when chromanol 293B is added - suggests Q1 channels switch
  • may be due to diffferent cell models or in vivo/ invitro
  • could switch only in some pateitns or at different stages of disease progression
  • second exp has camp stimulation but exp before does not have camp stimulation
22
Q

what is the K+ channel which causes K+ secretion on the apical membrane?

A
  • BK
23
Q

What can block apical K+ channels and what happens when this is applied?

A

paxilline

- when this is applied - less K+ secretion, lower liquid layer, 50% reduced cilia beat frequency

24
Q

what happens when artificial medium is added to BK KD cells?

A
  • brings liquid layer up to a normal height
  • cilia beat effectively
  • normal function
25
Q

what happens to Cl- secretion when apical BK channels are blocked by paxilline?

A

when apical BK channels are blocked, Cl- secretion is also blocked

  • shows that apical K+ channels also support Cl- secretion
  • needed for normal secretion of Cl- through CaCC
  • when BK KD less cl- secretion
26
Q

what is the final epithelial cell model which shows all Cl and K+ channels

A

apical - CFTR - Cl- secretion
apical - CaCC - Cl- secretion
apical - ENaC - Na absorption
apical - BK - K+ secretion
basolateral - hSK4 - K+ out of cell
basolateral - KvLQT1 (E3) - K+ out of cell
basolateral - Na/K ATPase - Na out of cell K+ into cell
basolateral - NKCC1 - Na, K, 2Cl into cell

epithelial (8 decks)

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