Insulin induces cation fluxes and increases intracellular calcium: DISCUSSION

The present study shows clearly that insulin causes a small inward current in HTC liver cells maintained near the rest­ing membrane potential in physiological cationic condi­tions. This insulin-induced current triggers a gradual depolarization of 5.7 mV. Such a slow depolarizing effect of insulin was observed previously following impalements of cultured rat hepatocytes with glass microelectrodes, but the underlying currents were not well defined. In the present study, we used ion substitutions to clarify the ionic compo­nents of the insulin-induced currents. In physiological cati- onic conditions, insulin-induced whole-cell currents have an average linear slope conductance of 663 pS and a reversal potential of-17.9 mV, located between the equilibrium po­tential for potassium ions, on one hand, and that for cations or chloride, on the other. Collapsing the potassium gradient brought the reversal potential of insulin-induced currents close to 0 mV, whereas moving Eq to +35 or -33 mV did not drive the reversal potential toward more positive or more negative values. In addition, no current was elicited by in­sulin administration when chloride ions were the major conducting ion species (all cations replaced by a nonper- meant species such as NMDG). These data strongly suggest that insulin-induced inward currents into HTC cells are mainly related to nonselective cationic components.

However, our results also suggest that insulin may induce a small parallel increase in potassium conductance as indi­cated by the reversal potential obtained in physiological cationic solutions (between Ek and Ecation). This conductive pathway may be related to the insulin-induced increase in intracellular calcium discussed below. Indeed, purinergic agonists such as ATP are known to trigger calcium-sensitive potassium currents in HTC cells, and calcium-activated potassium channels of small size have already been described in these cells by Lidofsky. Such channels could be a putative source of the potassium-selective component induced by insulin that drives the equilibrium potential of

insulin-induced currents partially away from 0 mV (nonselective cation equilibrium potential) in our physio­logical cationic conditions.

Nonetheless, our results indicate that the major action of insulin is to trigger inward cationic currents in HTC cells that consist principally of a nonselective component that probably contributes to the observed depolarization of HTC cell membrane potential and rise in cytosolic calcium.
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Indeed, in the present study, insulin was found to trigger an increase in HTC intracellular calcium that was generally monophasic and transient in nature. This corresponds to what we have recently described in primary rat hepatocytes. Similar to the latter study, the insulin-induced changes in hepatocellular calcium were absent when external cal­cium was chelated, in the present study. This effect was not related to altered insulin receptor occupancy, as confirmed by I insulin displacement studies, or to the depletion of internal calcium stores, as shown by the preserved response to the G-coupled purinergic receptor agonist ATP. These re­sults strongly suggest that insulin triggers an influx of exter­nal calcium into HTC cells in a manner similar to that found in primary hepatocytes.