Regulation of ion fluxes, cell volume and gap junctional coupling by cGMP in GFSHR-17 granulosa cells.
Gap junctional communication between granulosa cells seems to play a crucial role for follicular growth and atresia. Application of the double whole-cell patch-clamp- and ratiometric fura-2-techniques allowed a simultaneous measurement of gap junctional conductance ( G(j)) and cytoplasmic concentration of free Ca(2+) ([Ca(2+)](i)) in a rat granulosa cell line GFSHR-17. The voltage-dependent gating of G(j) varied for different cell pairs. One population exhibited a bell-shape dependence of G(j) on transjunctional voltage, which was strikingly similar to that of Cx43/Cx43 homotypic gap junction channels expressed in pairs of oocytes of Xenopus laevis. Within 15-20 min, gap junctional uncoupling occurred spontaneously, which was preceded by a sustained increase of [Ca(2+)](i) and accompanied by shrinkage of cellular volume. These responses to the whole-cell configuration were avoided by absence of extracellular Ca(2+), blockage of K(+) efflux, or addition of 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP) to the pipette solution. Even in the absence of extracellular Ca(2+) or blockage of K(+) efflux, formation of whole-cell configuration generated a Ca(2+) spike that could be suppressed by the presence of 8-Br-cGMP. We propose that intracellular cGMP regulates Ca(2+) release from intracellular Ca(2+) stores, which activates sustained Ca(2+) influx, K(+) efflux and cellular shrinkage. We discuss whether gap junctional conductance is directly affected by cGMP or by cellular shrinkage and whether gap junctional coupling and/or cell shrinkage is involved in the regulation of apoptotic/necrotic processes in granulosa cells.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects* / Q000502:physiology; D002450:Cell Communication / Q000187:drug effects* / Q000502:physiology; D048429:Cell Size / Q000187:drug effects; D002614:Chelating Agents / Q000494:pharmacology; D017630:Connexins / Q000378:metabolism; D006152:Cyclic GMP / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000502:physiology*; D003599:Cytoskeleton / Q000187:drug effects / Q000648:ultrastructure; D004533:Egtazic Acid / Q000494:pharmacology; D005260:Female; D016257:Fura-2 / Q000378:metabolism; D017629:Gap Junctions / Q000187:drug effects* / Q000502:physiology; D006107:Granulosa Cells / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D015640:Ion Channel Gating; D017136:Ion Transport / Q000187:drug effects* / Q000502:physiology; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D051381:Rats
D002614:Chelating Agents; D017630:Connexins; C016276:8-bromocyclic GMP; D004533:Egtazic Acid; D006152:Cyclic GMP; D011188:Potassium; D002118:Calcium; D016257:Fura-2