Lecture 10 - B subunits

Question 1

Beta subunits

Answer 1

are cytoplasmic/integral membrane proteins which regulate trafficking and Po

Question 2

KCNE family

Answer 2

vary in size 103-177aa
small mr
one TMD

Question 3

KCNQ1 changes it properties by…

Answer 3

depending on different Beta subunit effecting it

Question 4

Effect of KCNE1 on KCNQ1

Answer 4

E1 enhances Q1 function, the current is larger when both are expressed
Also reaches steady state faster

Question 5

KCNE1 and renal function

Answer 5

KCNE1 is on the apical membrane of the proximal tubule, important for glucose transport

Question 6

Proximal tubule proteins (3)

Answer 6

Apical = KCNE1
Basolateral = Na+/K+ATPase and K+/glucose cotransporter

Question 7

Clearance studies in vivo

Answer 7

anaesthetise mouse on heated pad, use rectal thermometer

cannulate carotid artery, jugular vein and ballder

Question 8

Why cannulate carotid artery?

Answer 8

BP measurement and blood samples

Question 9

Why cannulate jugular vein?

Answer 9

fluid replacement

Question 10

Why cannulate the bladder?

Answer 10

collecting urine for analysis

Question 11

E1 KO mice

Answer 11

no change in sodium or chloride plasma conc. decreased glucose conc. GFR normal

Question 12

Fractional excretion calculation

Answer 12

rate excretion/rate filtration

Question 13

E1 KO fractional excretion

Answer 13

sodium and chloride increase with water

no change in glucose

Question 14

E1 KO mice shows that…

Answer 14

E1 is most likely in the late proximal tubule therefore sodium channel is not working and causes a reduction in the sodium driving force

Question 15

Chromanol 293b

Answer 15

specific KCNQ1 inhibitor

Question 16

Chromanol 293b in WT

Answer 16

mimics an E1 KO

but no effect on KO

Question 17

Q1 KO clearance studies

Answer 17

no difference between KO and WT in sodium and water fractional excretion

Question 18

Bigger the chromosome-sensitive current…

Answer 18

greater channel function

Question 19

Patch clamp studies on E1 and Q1

Answer 19

WT does not show voltage-dependence so is not regulated by E1
therefore E1 does not regulate Q1

Question 20

Parietal cell basolateral membrane proteins

Answer 20

Na+/H+ exchanger
Na+/K+ATPase
Basolateral K+ channel
HCO3-/Cl- exchanger

Question 21

Parietal cell apical membrane proteins

Answer 21

chloride transporter
K+ channel
H+ATPase

Question 22

Parietal cell mechanisms

Answer 22

Co2 and H2O diffuse into the cell under carbonic anhydrase influence to produce HCO3- and H+
Chloride driving force for net secretion of HCL
ATP used to exchange K+ into the cell (recycled across the membrane) and H+ out to form HCL

Question 23

Parietal cell stimulants

Answer 23

acetylcholine
Histamine
Gastrin

Question 24

Acetylcholine stimulates…

Answer 24

M3

Question 25

Histamine stimulates…

Answer 25

H2

Question 26

Gastrin stimulates…

Answer 26

CCKB

Question 27

Ammonium pulse technique

Answer 27

ammonium added to EC solution, dissociates to NH3 and H+
NH3 enters cell and mops H+ up inside so pH increases
Take away EC ammonium so NH3 dissociates from H+ and moves out of cell, increased acidification in the cell

Question 28

Ammonium pulse technique shows

Answer 28

the rate of pH recovery of the cells to remove H+

stimulate carbochol/histamine then remove EC Na+ and measure rate in absence of a protein

Question 29

In H+/K+ATPase KO

Answer 29

no recovery seen

Question 30

KCNE2 KO

Answer 30

achlorhydic

pH is already very high under control conditions and does not react with added histamine

Question 31

KCNE2 KO ammonium pulse technique

Answer 31

histamine stimulated
KP secretes less H+
lack of function of K+/H+ATPase due to KCNE2 loss
apical membrane has Q1/E2 complex

Question 32

Q1 location

Answer 32

regulates KCNE2 ont he apical membrane of parietal cells