Regulation of the renal Na-HCO3 cotransporter: VII. Mechanism of the cholinergic stimulation.
Cholinergic agents regulate proximal tubule acidification but the mechanism responsible for this effect is unclear. We examined the effect of the cholinergic agent carbachol on the activity of the Na-HCO3 cotransporter in primary cultures of the proximal tubule of the rabbit. The activity of the cotransporter was assayed either as HCO3-dependent 22Na uptake or as the recovery of intracellular pH in cells perfused continuously with Cl-free physiologic solution containing amiloride to block the Na-H antiporter. Carbachol caused a dose-dependent stimulation of the cotransporter activity with a maximum increase of 90% above control values at 10(-5) M and half maximal stimulation at 10(-7) M. The stimulation was blocked by atropine and pirenzepine indicating an effect through the M1 muscarinic receptor. Carbachol increased intracellular calcium fourfold and the rise in cytosolic calcium was prevented by the intracellular calcium chelator, BAPTA. BAPTA also blocked the effect of carbachol on the cotransporter. Because carbachol activates phospholipase C and protein kinase C, we examined the effect of carbachol in the presence of the phospholipase C inhibitor, U73122, or the PKC inhibitor, calphostin C, or PKC depletion. The phospholipase C inhibitor prevented both the effect of carbachol on the cotransporter and on the intracellular Ca. Calphostin C and PKC depletion also prevented the stimulation of the cotransporter. Carbachol increased PKC activity and caused translocation of the PKC to the particulate fraction. We also examined the effect of the phosphatase inhibitor, calyculin A or the calmodulin kinase inhibitor, W-13 on carbachol stimulation. Calyculin A and W13 likewise prevented the carbachol-induced stimulation of the cotransporter. These results demonstrate that cholinergic stimulation modulated the activity of the cotransporter through multiple pathways including phospholipase C/PKC and phosphatase systems.
Ruiz
O S
OS
Department of Medicine, University of Illinois at Chicago, 60612-7315, USA.
Qiu
Y Y
YY
Cardoso
L R
LR
Arruda
J A
JA
D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D018678:Cholinergic Agents / Q000494:pharmacology*; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D017136:Ion Transport / Q000187:drug effects; D007687:Kidney Tubules, Proximal / Q000187:drug effects* / Q000378:metabolism*; D008387:Marine Toxins; D009281:Naphthalenes / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D011817:Rabbits; D012964:Sodium / Q000378:metabolism*; D027982:Sodium-Bicarbonate Symporters; D013449:Sulfonamides / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism
D001639:Bicarbonates; D002352:Carrier Proteins; D002614:Chelating Agents; D018678:Cholinergic Agents; D004791:Enzyme Inhibitors; D004963:Estrenes; D008387:Marine Toxins; D009281:Naphthalenes; D010080:Oxazoles; D011760:Pyrrolidinones; D027982:Sodium-Bicarbonate Symporters; D013449:Sulfonamides; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D004533:Egtazic Acid; C059041:calyculin A; C033772:N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide; D002217:Carbachol; D012964:Sodium; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D010738:Type C Phospholipases; C058819:calphostin C; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium