Impaired inhibitory G-protein function contributes to increased calcium currents in rats with diabetic neuropathy

There is a growing body of evidence that sensory neuropathy in diabetes is associated with abnormal calcium signaling in dorsal root ganglion (DRG) ne urons. Enhanced influx of calcium via multiple high-threshold calcium curre nts is present in sensory neurons of several models of diabetes mellitus, i ncluding the spontaneously diabetic BioBred/Worchester (BB/W) rat and the c hemical streptozotocin (STZ)-induced rat. We believe that abnormal calcium signaling in diabetes has pathologic significance as elevation of calcium i nflux and cytosolic calcium release has been implicated in other neurodegen erative conditions characterized by neuronal dysfunction and death. Using e lectrophysiologic and pharmacologic techniques, the present study provides evidence that significant impairment of G-protein-coupled modulation of cal cium channel function may underlie the enhanced calcium entry in diabetes. N- and P-type voltage-activated, high-threshold calcium channels in DRGs ar e coupled to mu opiate receptors via inhibitory G(o)-type G proteins. The r esponsiveness of this receptor coupled model was tested in dorsal root gang lion (DRG) neurons from spontaneously-diabetic BB/W rats, and streptozotoci n-induced (STZ) diabetic rats. Intracellular dialysis with GTP gammaS decre ased calcium current amplitude in diabetic BB/W DRG neurons compared with t hose of age-matched, nondiabetic controls, suggesting that inhibitory G-pro tein activity was diminished in diabetes, resulting in larger calcium curre nts. Facilitation of calcium current density (I-DCa) by large-amplitude dep olarizing prepulses (proposed to transiently inactivate G proteins), was si gnificantly less effective in neurons from BB/W and STZ-induced diabetic DR Gs. Facilitation was enhanced by intracellular dialysis with GTP gammaS, de creased by pertussis toxin, and abolished by GDP betaS within 5 min. Direct measurement of GTPase activity using opiate-mediated GTP gamma[S-35] bindi ng, confirmed that G-protein activity was significantly diminished in STZ-i nduced diabetic neurons compared with age-matched nondiabetic controls. Dia betes did not alter the level of expression of mu opiate receptors and G-pr otein alpha subunits. These studies indicate that impaired regulation of ca lcium channels by G proteins is an important mechanism contributing to enha nced calcium influx in diabetes.