Site and mechanism of action of trichlormethiazide in rabbit distal nephron segments perfused in vitro.

Abstract

To determine the exact site and mechanism of action of thiazide diuretics, effects of 104 M trichlormethiazide (TCM) on NaCJ transport were examined in the distal convoluted tubule (DCT), the connecting tubule (CNT) and the cortical collecting duct (CCD) of rabbit kidney by the in vitro microperfusion technique. TCM added to the lumen decreased lumen-to-bath 'Cl flux (JC(LB)) only in the CNT without changing the transmural voltage (VT). In the DCT, 10-4 M furosemide did not change JCI(LB) even if it was added to the lumen with 10-4 M TCM, whereas 10-5 M amiloride in the lumen decreased the lumen-to-bath 22Na flux (JNa(LB)) and VT. In the CNT, TCM added to the lumen did not affect the bath-to-lumen OC1 flux. Addition of TCM to the bath slightly decreased JC(LB). Luminal addition of 10-4 M TCM also decreased JNa(LB). Amiloride at 10-5 M in the lumen decreased both JNa(LB) and VT.Addition of TCM with 10-5 M amiloride further decreased JNa(LB) without affecting VT, indicating that TCM affects the electroneutral Na+ transport, which is distinct from the amiloride-sensitive conductive Na' pathway. When Na+ was removed from the lumen, JCI(LB) was markedly decreased, but addition of TCM did not cause further decrease in JCI(LB). Furosemide did not affect JC(LB), but addition of both 10-4 M TCM and furosemide decreased JCI(LB), indicating that Na+-K+-2CI-cotransport is not involved in the action of TCM.Removal of HCO3 slightly decreased JCI(LB), and TCM caused further decrease in JCI(LB). Amiloride at 10-3 M, a concentration supposed to inhibit the Na+/H+ antiport, slightly decreased JCI(LB), and addition of TCM caused a further marked decrease in JJI(LB). The similar results were also obtained when the combined effects of 10-3 M 4,4'-diisothiocyano-stilben-2,2'-disulfonate(DIDS) and 10'-M TCM were examined.These findings suggest that the parallel antiport of Na+/H+ and Cl-/HCO-is not involved in the action of TCM. By excluding other possible mechanisms involving neutral Na+-dependent Cl-transport, we conclude that TCM inhibits Na+-ClI cotransport in the luminal membrane of the rabbit CNT. IntroduptionAlthough thiazide diuretics have been widely used in the treatment of edema and hypertension for many years, the exact Address reprint requests to Dr. Imai,

Effect of S-8666 on Cl- Transport in the Rabbit Connecting Tubule Perfused in Vitro.

“…Thus, relative hypocalciuric effect of S-8666 (Shimizu et al 1989a) may be explained also by its action on the CNT. Since we have also demonstrated that furosemide did not affect Cl-transport in the CNT (Shimizu et al 1988c), the additive diuretic effect of S-8666 and furosemide (Shimizu et al 1989b) may be accounted for by the action of S-8666 on the CNT.…”

“…In the previous study (Shimizu et al 1988c), we have established that in the rabbit thiazide diuretics inhibit Na+-C1-cotransport by acting exclusively on the ONT. Neither distal convoluted tubule (DOT) nor cortical collecting duct (COD) is the target of the action of thiazide diuretics.…”

“…S-8666 at 1 mM suppressed C1-flux by 15%, whereas 0.1 mM TCM decreased it by 28% (Shimizu et al 1988c). It appears that the potency of S-8666 in inhibiting Nat-Cl cotransport in the CNT is much less than that of TCM, although the available data do not allow us to make exact comparison.…”

“…The rabbit CNTs were perfused in vitro as reported previously (Imai 1979 ;Shimizu et al 1988c ;Yoshitomi et al 1988 ;Muto et al 1990). The CNT was identified according to the criteria reported previously (Imai 1979).…”

“…Since anticalciuric effect is the most unique for thiazide diuretics, we speculated that an additional site of action of S-8666 might be common to that of thiazide diuretics. Shimizu et al (1988c) reported that in the rabbit trichlormethiazide (TCM) inhibits Nat-Cl cotransport in the connecting tubule (CNT). Therefore, we examined effects of S-8666 on 01 -transport, transmural voltage (VT), and basolateral membrane voltage (VB) in the rabbit CNT to provide direct evidence in support of the view that an additional site of action of S-8666 is common to that of thiazide diuretics.…”