Beta subunit interactions Flashcards
What are beta subunits, examples of some
They are proteins that dont have a role in ion channels- they modify the properties of the alpha subunits- which do act as ion channels
KCNE family- 1,2 and 3
Properties of the KCNE family
1,2 and 3 are found in the epithelia vary in molecular weight 1 TMSD E1 is mutated in long QT syndrome 1,2 and 3 have the ability to regulate the Q1 K channel
Whats the effect of E1 on Q1- how was this shown?
Xenopus occytes- expressing cRNA encoding KCNQ1 alone or with E1
two-electrode voltage clamp
With E1= larger currents, slower activation times, shifted voltage dependence (but still there)
What did expression studies of E1 and Q1 highlight?
E1 is expressed in the kidney- the majority of it is found in the proximal tubule
found in the apical membrane
Q1 is expressed in the apical membrane in the kidney, but mainly in the distal kidney
only a small bit of overlap
How are clearance studies carried out?
In mice
cannulate bladder- analyse ion concentrations and volume of urine
cannulate carotid artery- take BP measurements and analyse plasma ion concentrations
cannulate jugular vein- fluid replacement
do this in WT and KO mice
Was there a change in plasma concentrations of ions in the E1 KO mice?
Maintain normal Na and Cl plasma conc
Glucose conc was significantly lower in the KO mice- not reabsorbing enough glucose
*but- plasma glucose conc was abnormally elevated in the WT and KO mice
Was there a change in fractional excretion in the E1 KO mice?
FE of Na, Cl, glucose and fluid was higher
struggling to reabsorb these ions
glucose results might explain why the plasma glucose was lower
However in another study- no change in the FE of glucose- which makes more sense, E1 has a role in the later parts of the proximal tubule, which tend not to be important in reabsorbing glucose
What does a change in FE indicate and what seems to be going on in the E1 KO mice?
Changes in FE are indicative of a change in tubular function, losing E1 is impacting on the nephron- defect somewhere
If you lose E1- lose contribution to the resting membrane potential- reduce the driving force for Na reabsorption- this then reduces the the absorption of chloride ions- and therefore fluid as well
How did 293B effect the FE of NA in WT and E1 KO mice?
WT mice- increased the FE of Na significantly- from 2-4% approx
KO mice- no effect- Q1 channels aren’t working in these mice- nothing to block
Chromanol turns WT FE to KO FE
E1 is regulating a chromanol sensitive channel
Was there a change in fractional excretion in Q1 KO mice?
Slightly reduced FE of sodium and water - but NOT significant
-the main role of E1 does not seem to be regulating Q1 K channels in the kidney
How did they measure whole cell current and what did they conclude?
Dissected out the late proximal tubule- used the patch clamp technique to measure total whole cell current- then added 293B and measured how much the current went down by
Realised that the 293B sensitive current is not the same as the voltage-dependent Q1/E1 current they predicted
didnt show voltage dependence
conclusion: E1 is regulating a K channel that isnt Q1
How is HCL secreted by the parietal cells in the stomach?
Apical Cl channel- secretes Cl into the stomach- Cl gradient generated by the chloride-bicarbonate exchanger= bicarb out, chloride in
Secretion of H ions through a H/K ATPase- secretes H into the stomach for the exchange of potassium- there isnt enough potassium in the stomach to sustain this-
potassium comes from an apical potassium channel- potassium released then goes through the H/K ATPase
What is the ammonium pulse technique?
Ammonia is added to the cell- goes into the cell and combines with H to form NH4+- see an increase in pH in this first phase
then ammonia is taken away- ammonium breaks apart and H is released (very quick)- massive increase in H+ concentration= acidification and the pH goes down
pH of the cell then recovers- measure of how fast H+ ions are secreted
How do Q1 KO cells respond to an acid load?
no pH recovery at all
K channel in apical membrane doesnt work- so no K goes into the cell in exchange for H ions- no H secretion
How do E2 KO cells respond to an acid load?
no pH recovery at all- H/K ATPase doesnt work
