Earlier, we showed that streptozocin (STZ)-induced type 1 diabetes in rats leads to the development of painful peripheral diabetic neuropathy (PDN) manifested as thermal hyperalgesia and mechanical allodynia accompanied by significant enhancement of T-type calcium currents (T-currents) and cellular excitability in medium-sized dorsal root ganglion (DRG) neurons. Here, we studied the in-vivo and in-vitro effects of gene-silencing therapy specific for the Cav3.2 isoform of T-channels, on thermal and mechanical hypersensitivity, and T-current expression in small and medium-size DRG neurons of STZ-treated rats. We found that silencing of the T-channel Cav3.2 isoform using antisense oligonucleotides, had a profound and selective anti-hyperalgesic effect in diabetic rats and is accompanied by significant down-regulation of T-currents in DRG neurons. Anti-hyperalgesic effects of CaV3.2 antisense oligonucleotides in diabetic rats were similar in models of rapid and slow onset of hyperglycemia following intravenous and intraperitoneal injections of STZ, respectively. Furthermore, treatments of diabetic rats with daily insulin injections reversed T-current alterations in DRG neurons in parallel with reversal of thermal and mechanical hypersensitivity in-vivo. This confirms that CaV3.2 T-channels, important signal amplifiers in peripheral sensory neurons, may contribute to the cellular hyperexcitability that ultimately leads to the development of painful PDN.

OBJECTIVEMorbid obesity may be accompanied by diabetes and painful diabetic neuropathy, a poorly understood condition that is manifested by mechanical or thermal allodynia and hyperalgesia. Recent studies have highlighted the importance of T-type calcium channels (T-channels) in peripheral nociception; therefore, our goal was to examine the function of these channels in the pathophysiology and development of painful diabetic neuropathy.RESEARCH DESIGN AND METHODSIn vivo testing of mechanical and thermal sensation, morphometric peripheral nerve studies, and electrophysiological and biochemical measurements were used to characterize the role of T-channels and the development of painful diabetic neuropathy in leptin-deficient (ob/ob) mice.RESULTSWe found that ob/ob mice developed significant mechanical and thermal hypersensitivity early in life that coincided with hyperglycemia and was readily reversed with insulin therapy. These disturbances were accompanied by significant biophysical and biochemical modulation of T-channels in dorsal root ganglion neurons as measured by a large increase in the amplitude of T-currents and the expression of mRNA. The most prevalent subtype, α1H (Cav3.2), was most strongly affected. Moreover, (3β,5α,17β)-17-hydroxyestrane-3-carbonitrile (ECN), a novel neuroactive steroid and selective T-channel antagonist, provided dose-dependent alleviation of neuropathic thermal and mechanical hypersensitivity in diabetic ob/ob mice.CONCLUSIONSOur results indicate that pharmacological antagonism of T-channels is potentially an important novel therapeutic approach for the management of painful diabetic neuropathy.

General anesthetics, either alone or in combination, can be detrimental to the developing mammalian brain and induce extensive apoptotic degeneration of immature neurons when they are administered at the peak of synaptogenesis. Because neuron development and normal functions depend on the integrity and astroglia, we sought to determine whether general anesthesia also causes disturbances in the early development of astroglia. Using isoflurane, an inhaled anesthetic that is highly toxic to immature neurons, we studied primary astroglia cultures, focusing on very early development (day-in-vitro 4 treatment). Exposure to 3% isoflurane for 24 hours delayed morphological differentiation and impaired the growth of the immature astrocytes. The timing of delayed astroglia maturation and growth coincided with a major disturbance in actin cytoskeleton sculpting that was manifest as impaired actin stress fiber formation and cytoskeletal organization and downregulation of the focal adhesion protein, paxillin. Isoflurane-induced actin cytoskeletal changes were accompanied by a significant decrease in protein levels of the endogenous GTPase RhoA that regulates the phosphorylation of myosin light chain protein, suggesting that isoflurane-induced impairment in glial growth and morphological development is, in part, mediated by the RhoA/myosin light chain protein signaling pathway.

α-Lipoic acid (1,2-dithiolane-3-pentanoic acid; lipoic acid) is an endogenous compound used to treat pain disorders in humans, but its mechanisms of analgesic action are not well understood. Here we show that lipoic acid selectively inhibited native CaV3.2 T-type calcium currents (T-currents) and diminished T-channel-dependent cellular excitability in acutely isolated rat sensory neurons. Lipoic acid locally injected into peripheral receptive fields of pain-sensing sensory neurons (nociceptors) in vivo decreased sensitivity to noxious thermal and mechanical stimuli in wild-type but not CaV3.2 knockout mice. Ensuing molecular studies demonstrated that lipoic acid inhibited recombinant CaV3.2 channels heterologously expressed in human embryonic kidney 293 cells by oxidating specific thiol residues on the cytoplasmic face of the channel. This study provides the first mechanistic demonstration of a nociceptive ion channel modulation that may contribute to the documented analgesic properties of lipoic acid in vivo.