Dorsal root ganglion application of muscimol prevents hyperalgesia and stimulates myelin protein expression after sciatic nerve injury in rats.

BACKGROUND: Peripheral nerve injuries may result in debilitating pain that is poorly responsive to conventional treatment. Neuropathic pain induced by peripheral nerve injury is caused, in part, by ectopic discharges from the injury site or the dorsal root ganglia (DRG) resulting in enhanced central input and central hyperexcitability. A heterogeneous family of γ-aminobutyric acid (GABA)(A) channels is important in quieting neuronal excitability. We have recently reported that in vivo modulation of GABAergic neurons in DRG can alter the course of neuropathic pain development after peripheral nerve injury. It seems that direct application of a potent GABA(A) agonist, muscimol, to the ipsilateral DRG prevents the development of hyperalgesia in rats subjected to a sciatic nerve crush injury. In addition to potentially curtailing hyperexcitability, GABAergic stimulation upregulated expression of peripheral myelin protein 22 (PMP22), a key component of the basal lamina. PMP22 expression correlates with peripheral myelin formation and nerve regeneration. METHODS: Because of the importance of PMP22 for the formation and stability of myelin, and the fact that PMP22 expression could be GABAergically modulated, we examined whether direct DRG application of muscimol can restore PMP22 protein expression and the integrity of nerve fibers after crush injury of a sciatic nerve. RESULTS: Using adult female rats and a crush injury model, we found that GABAergic modulation in the ipsilateral DRG restores PMP22 protein expression in the distal segment of the sciatic nerve and improves myelin stability in the basal membrane of nerve fibers, thus giving the morphological appearance of lessened nerve injury or faster nerve fiber regeneration. Both the enhanced PMP22 protein expression and morphological improvements coincide with the abolishment of thermal and mechanical hypersensitivity. CONCLUSIONS: The DRG could be a promising therapeutic target in nerve regeneration and pain alleviation after crush injury of a myelinated peripheral nerve.

Isoflurane impairs immature astroglia development in vitro: the role of actin cytoskeleton.

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 of 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 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.

Selective T-type calcium channel blockade alleviates hyperalgesia in ob/ob mice.

OBJECTIVE: Morbid 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 METHODS: In 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. RESULTS: We 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, alpha1H (Ca(v)3.2), was most strongly affected. Moreover, (3beta,5alpha,17beta)-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. CONCLUSIONS: Our results indicate that pharmacological antagonism of T-channels is potentially an important novel therapeutic approach for the management of painful diabetic neuropathy.

In vivo silencing of the Ca(V)3.2 T-type calcium channels in sensory neurons alleviates hyperalgesia in rats with streptozocin-induced diabetic neuropathy.

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 Ca(V)3.2 isoform of T-channels, on thermal and mechanical hypersensitivities, and T-current expression in small- and medium-sized DRG neurons of STZ-treated rats. We found that silencing of the T-channel Ca(V)3.2 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 Ca(V)3.2 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 hypersensitivities in vivo. This confirms that Ca(V)3.2 T-channels, important signal amplifiers in peripheral sensory neurons, may contribute to the cellular hyperexcitability that ultimately leads to the development of painful PDN.