Effect of glandular kallikrein on distal bicarbonate transport. Role of basolateral Cl-/HCO3- exchanger and vacuolar H+-ATPase

The luminal membrane of collecting duct cells, specially the intercalated c ells, is normally exposed to active kallikrein. This is due to the specific localization of renal kallikrein in the connecting tubule cells. We have p reviously reported inhibition of distal bicarbonate secretion by renal kall ikrein. The present study was performed to evaluate the participation of basolatera l Cl-/HCO3- exchanger and luminal H+-ATPase activity of cortical collecting duct segments (CCD) in the mechanism involved in the inhibition of bicarbo nate secretion induced by the enzyme. The effect of orthograde injections o f 1 mu g/ml (250U/6.3mg) pig pancreatic kallikrein, in the absence and pres ence of 1 mM DIDS (stilbene-disulfonic acid) in the renal tubule system, wa s evaluated. Urine fractions were collected after two-minutes stop-flow. Ch anges in the urine fraction (Fr) related to those in free-flow urine sample s (Ff) were related to the respective polyfructosan (Inutest) ratio. Renal kallikrein activity (Fr:Ff kallikrein/Fr:Ff polyfructosan) increased signif icantly in the first 120 mu l urine fraction collected after glandular 1 mu g/ml kallikrein, P<0.05, (first stop-flow) and after glandular 1 mu g/ml k allikrein plus 1 mM. DIDS P<0.05 (second stop flow). Bicarbonate secretion rate (Fr:Ff HCO3-/ Fr:Ff polyfructosan) of collecting ducts was significant ly reduced in the first 120 mu l urine fraction collected related to contro l, during the first and second stop-flow periods. No difference was shown i n bicarbonate excretion between the first 120 mu l urine fractions collecte d after administration of glandular kallikrein and glandular kallikrein plu s DIDS. To measure H+-ATPase activity, rat microdissected cortical collector tubule s (CCD) were incubated in the presence of increasing glandular kallikrein d oses (A: 93, B: 187 and C: 375 mU/200 mu L) in the presence of ouabain (4 m u M) and omeprazole (100 mu M) to inhibit Na+- K+-ATPase and H+- K+-ATPase, respectively. In CCD, bafilomycin-sensitive H+-ATPase activity (pmol/mm/mi n) after increasing kallikrein doses did not differ significantly from cont rol. No difference related to control H+-ATPase activity was observed when microdissected CCD segments were incubated in the presence of an AT, recept or antagonist (Losartan 10(-6) M) and glandular kallikrein(93 mU). On the c ontrary, angiotensin II (10(-8) M) significantly decreased H+-ATPase activi ty. The present study shows that neither basolateral Cl- /HCO3- exchanger nor H +-ATPase activity are involved in bicarbonate inhibition by glandular kalli krein at CCD. Involvement of luminal Cl-/HCO3- exchanger at beta intercalat ed cells in CCD may be suggested for the bicarbonate secretion inhibition i nduced by renal kallikrein.