Regulatory effects of insulin-like growth factor-1 on the expression of sensory neuropeptide mRNAs in cultured dorsal root ganglion neurons with excitotoxicity induced by glutamate.

OBJECTIVE: To determine the effects of insulin-like growth factor-1 (IGF-1) on the expression of preprotachykinin (PPT) mRNA encoding substance P (SP) and calcitonin gene-related peptide (CGRP) mRNA in cultured dorsal root ganglion (DRG) neurons with excitotoxicity induced by glutamate (Glu). METHODS: DRGs were dissected from embryonic day 15 Wistar rats. DRG neurons were dissociated and cultured for 48 h and then exposed to Glu (0.2 mmol/L) or Glu (0.2 mmol/L) plus IGF-1 (5 nmol/L, 10 nmol/L and 20 nmol/L) for 12 h. The DRG neurons in control group were exposed to only growth media throughout the experiment. After that, the living DRG neurons were observed under inverted phase contrast microscope and microphotographs were taken. The expression levels of PPT and CGRP mRNAs were detected by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: IGF-1 could inhibit Glu-induced shortening of neurite. Besides, IGF-1 could significantly increase the levels of PPT mRNA and CGRP mRNA in primary cultured DRG neurons with Glu-induced excitotoxicity, in a dose-dependent manner. CONCLUSION: IGF-1 may exert neuroprotective effects on DRG neurons against Glu-induced excitotoxicity, probably through regulating the expression levels of PPT and CGRP mRNAs.

Butyrate sensitizes the release of substance P and calcitonin gene-related peptide evoked by capsaicin from primary cultured rat dorsal root ganglion neurons.

OBJECTIVES: To investigate whether butyrate increases substance P (SP) and calcitonin gene-related peptide (CGRP) release evoked by capsaicin from primary cultured dorsal root ganglion (DRG) neurons. METHODS: DRG was dissected out from embryonic 15-day-old Wistar rat and cultured as dissociate cells for 24 h then exposed to butyrate (0.01 mmol/L, 0.1 mmol/L, 1 mmol/L, 10 mmol/L, respectively) for another 48 h. The neurons cultured continuously in media served as normal control. All above cultured samples were processed for detecting expression of mRNA for SP, CGRP and vanilloid receptor 1 (VR1) of DRG neurons by RT-PCR, and VR1 protein expression by Western blot. SP and CGRP basal release levels were measured by radioimmunoassay (RIA). After that, the DRG cells for RIA were stimulated by capsaicin (300 nmol/L) for 5 min and the culture media were harvested for detecting SP and CGRP release levels by RIA. The neurons exposed to vehicle solution served as vehicle controls. RESULTS: Exposure of butyrate on DRG neurons at higher concentrations (1 mmol/L, 10 mmol/L) for 48 h increased expression mRNA for SP and CGRP than that at lower concentrations (0.01 mmol/L, 0.1 mmol/L) and normal control (P<0.001). VR1 mRNA and VR1 protein expression increased in a dose-dependent manner after exposure of different concentrations of butyrate. Butyrate did not alter the basal release, significantly enhanced neuropeptide release evoked by capsaicin. SP and CGRP release levels in the culture media exposed by butyrate at higher concentrations (1 mmol/L, 10 mmol/L) for 48 h and then stimulated by capsaicin were higher than that at lower concentrations (0.01 mmol/L, 0.1 mmol/L) (P<0.001). The exposure of vehicle solution did not produce any increase of SP and CGRP release from primary cultured DRG neurons. DISCUSSION: Butyrate may promote the expression of mRNA for SP, CGRP and increase sensitivity of capsaicin on SP and CGRP release from primary cultured rat dorsal root ganglion neurons. The promotion of VR1 mRNA and VR1 protein expression by butyrate implicated that VR1 may be involved in the mechanisms of sensory neuropeptide release evoked by capsaicin.