Histochemical, biochemical & electrophysiological studies on dorsal root ganglion cultures : Focusing on galanin, neuropeptide Y & glutamate

Galanin (Gal) and neuropeptide Y (NPY) are two peptides, which are strongly up- regulated in a population of dorsal root ganglion (DRG) neurons after peripheral nerve injury (axotomy). Both peptides may be involved in the control of pain sensation, especially after nerve injury (neuropathic pain). The main transmitter in DRG neurons is, however, the excitatory amino acid glutamate (Glu), while neuropeptides may have multiple roles including modulation and trophic influences. The aim of this thesis work was to characterize dispersed DRG neurons in culture, a putative model of axotomy, by (1) monitoring Gal and NPY and their transcripts, (2) investigating the effects of neurotrophic factors on peptide expression, Glu release and calcium levels and, finally, (3) studying the effect of exogenously applied Gal on cultured DRG neurons with electrophysiological methodology. We have shown that expression of Gal, NPY and their transcripts is similar in long-term cultures as in DRGs in vivo after axotomy. The remarkable increase of Gal level in cultures could be counteracted mainly by leukemia inhibitory factor (LIF) and basic fibroblast growth factor (bFGF). LIF was also shown to be involved in the local regulation of regeneration. The high NPY levels in DRG cultures could not be counteracted with any of several trophic factors. Instead, acidic FGF and bFGF further increased NPY expression. Moreover, the nonapeptide bradykinin induced a marked increase of Glu release, which was attenuated by NGF and BDNF and, in fact, completely counteracted by bFGF Finally, in electrophysiological studies Gal significantly increased the excitability and the peak Ca2+ currents of small size DRG neurons in acutely dissociated cultures. The latter effect is reduced both in preaxotomized and long term-cultured neurons