Short term regulation of cell-cell communication in TM3 Leydig cells. A perforated patch study

Determination of the junctional conductance (g(j)) in TM3 Leydig cells by the dual whole cell patch clamp technique (DWCPC) shows that coupling undergoes a rapid and irreversible run down. Addition of ATP or cAMP derivatives to the pipette solution has been shown to prevent this phenomenon in several tissues, but this same treatment is unable to inhibit run down in Leydig cells. Because the run down in junctional conductance may pose serious problems to the interpretation of results, we also measured g(j) by using the double perforated patch clamp technique (DPPT). Access to the cell interior was achieved by adding 200 microgram/ml of nystatin to the pipette solution. With this method, run down in g(j) was greatly reduced, amounting to no more than 5% of the initial value. Exposure of the cells, under DWCPC or DPPT, to dibutyryl cAMP or to tumor promoting agent (TPA) led to a decrease in cell to cell communication. Staurosporine, a PKC inhibitor, increased g(j) and was able to prevent and reverse the uncoupling action of cAMP or TPA. Our results indicate that cell-cell communication in Leydig cells is down regulated by both protein kinases A and C, interacting in a complex manner.