Pancreatic acinar cells: the acetylcholine equilibrium potential and its ionic dependency

Abstract

Two glass micro-electrodes were inserted into neighboring cells from rat or mouse pancreatic segments, superfused in vitro. The tip of a 3rd glass micro-electrode, filled with 2 M-AChCl, was placed just outside the acinus under investigation. Membrane potential and resistance, and changes in these parameters in response to short pulses of ACh stimulation, were recorded. The resting current-voltage relationship, obtained by injecting 100 ms depolarizing or hyperpolarizing current pulses through 1 of the intracellular micro-electrodes and recording the membrane potential with the other intracellular electrode, was linear within the range -5 to -60 mV. Injecting depolarizing or hyperpolarizing current (d.c.) through 1 of the intracellular micro-electrodes, the membrane potential (as measured with the other intracellular micro-electrode) could be set at various levels. The ACh effect at different membrane potentials was investigated. When acinar cell membrane was hyperpolarized, the amplitude of ACh-evoked depolarization was increased, while ACh-evoked depolarization was reduced when the membrane potential was reduced by depolarizing current and finally changed into a hyperpolarization at very low membrane potentials. In each acinus investigated (rat and mouse), there was a linear relationship between amplitude of ACh-evoked potential change (.DELTA.V) (+ value or - value according to polarity) and resting membrane potential. During superfusion with control solution, the value of the membrane potential at which ACh did not evoke a potential change (EACh) was about - 15 mV in the mouse and about -20 mV in the rat. During superfusion with a Cl-free sulfate-containing solution (steady state), a linear relationship between .DELTA.V and resting membrane potential was again found, but EACh (mouse) was about +10 mV. A continuous rough estimate of EACh was obtained by injecting repetitively depolarizing current pulses (100 ms) through 1 intracellular micro-electrode; in this way, the effect of ACh measured by the other intracellular electrode could be assessed simultaneously at the spontaneous resting level and at a depolarized level. The direction of change in EACh following acute changes in the superfusion fluid ion composition was assessed. Replacing extracellular Cl- by sulfate caused an immediate change in EACh in the positive direction. Cl- readmission after a long period of Cl- ion deprivation caused an immediate sharp change in EACh in the negative direction. Replacing extracellular Na+ by Tris caused an immediate transient negative change in EACh. Taking away extracellular Ca2+ changed EACh in a positive direction. Augmenting extracellular K+ concentration to 40 mM caused a change in EACh in the positive direction. At a membrane potential (V) equal to EACh the sum of ionic currents evoked by ACh action was zero. Using the Goldman treatment, ACh possibly increased membrane Na+, K+ and Cl- permeability. The approximate relative ion permeabilties of the pathways opened up by ACh are: P[permeability]Na/PK = 2.5 and PCl/PK = 5. At V = EACh, the approximate relative sizes of the ACh-evoked currents are: I[influx]Na/IK = 2.6 and ICl/IK = 1.6. ACh caused Na+ and Cl- influx and a small K+ efflux.