Carbachol activates a K⁺ channel of very small conductance in the basolateral membrane of rat pancreatic acinar cells

Abstract.

Secretion of Cl^– requires the presence of a K⁺ conductance to hyperpolarize the cell, and to provide the driving force for Cl^– exit via luminal Cl^– channels. In the exocrine pancreas Cl^– secretion is mediated by an increase in cytosolic Ca²⁺ ([Ca²⁺]_i). Two types of Ca²⁺-activated K⁺ channels could be shown in pancreatic acinar cells of different species. However, there are no data on Ca²⁺-activated K⁺ channels in rat pancreatic acini. Here we examine the basolateral K⁺ conductance of freshly isolated rat pancreatic acinar cells in cell-attached and cell-excised patch-clamp experiments. Addition of carbachol (CCH, 1 µmol/l) to the bath led to the activation of very small conductance K⁺ channels in cell-attached patches (n=27), producing a noisy macroscopic outward current. The respective outward conductance increased significantly by a factor of 2.1±0.1 (n=27). Noise analysis revealed a Lorentzian noise component with a corner frequency (f _c) of 30.3±3.5 Hz (n=19), consistent with channel activity in these patches. The estimated single-channel conductance was 1.5±0.4 pS (n=19). In cell-excised patches (inside out) from cells previously stimulated with CCH, channel activity was only observed in the presence of K⁺ in the bath solution. Under these conditions f _c was 47.6±11.9 Hz (estimated single-channel conductance 1.1±0.2 pS, n=20). The current/voltage relationship of the noise showed weak inward rectification and the reversal potential shifted towards E _K ⁺ when Na⁺ in the bath was replaced by K⁺. Channel activity in cell-excised patches was slightly reduced by 10 mmol/l Ba²⁺ (23.6±2.1% of the total outward current) and was completely absent when K⁺ in the bath was replaced by Na⁺. Reduction of the [Ca²⁺]_i from 1 mmol/l to 1 µmol/l in cell-excised experiments decreased the current by 52.3±12.3% (n=5). Expression of K_VLQT1, one of the possible candidates for a small-conductance K⁺ channel in rat pancreatic acinar cells, was shown by reverse transcriptase polymerase chain reaction (RT-PCR). In fact, a K_VLQT-blocker (chromanol 293B) reduced channel activity in seven excised patches. These data suggest that CCH activates very small conductance K⁺ channels in rat pancreatic acinar cells, most likely via an increase in [Ca²⁺]_i.