The effects of a range of neuronal excitants were examined on the firing of central neurones of the cerebral cortex, ventrobasal thalamus, dentate gyrus and dorsal and ventral horns of the spinal cords of urethane anaesthetized rats. These responses were pharmacologically characterized on the basis of their susceptibilities to a number of antagonists and from these results, inferences were made concerning probable receptor mechanisms employed by the agonists. Throughout these experiments the technique of iontophoresis was found to be an ideal one for evaluating the effects of agonists and antagonists on single neurones. Neurones in the cortex, thalamus and Renshaw cells of the spinal cord were readily excited by acetylcholine. These responses were elicited also by both nicotinic and muscarinic cholinomimetics. Excitations produced by acetylcholine and acetyl-β-methylcholine were antagonized by atropine and those of acetylcholine and nicotinic agonists were blocked by nicotinic antagonists. The results may be interpreted as revealing a difference between excitatory cholinergic receptors in the rat and in the cat; the nature of these receptors is discussed. to The excitatory responses of ventrobasal thalamic neurones iontophoretically applied amino acids related to glutamate and aspartate could be blocked both by glutamate diethylester and α-aminoadipate. These two antagonists were found to possess different mechanisms of action however, as the ranking orders of susceptibility of the agonists differed for each antagonist. An analysis of these orders led to the proposal that more than one and possibly as many as three different receptors for the excitatory amino acids exist on central neurones. A number of additional compounds were tested for an evaluation of their antagonistic properties against the amino acid induced responses, and these results are discussed in light of possible steric requirements of the receptors. Granule cells of the dentate gyrus were excited by the amino acids and by their synaptic responses to stimulation of perforant path and commissural inputs. A differential effectiveness of glutamate diethylester and α-aminoadipate was suggestive that two distinct excitatory amino acid receptors, both of which appear to be of synaptic significance, coexist on the same neurones. The effects of octopamine were compared with those of catecholamines on neurones of the cortex and dorsal horn of the spinal cord. Both excitation and depression of neuronal firing was observed with octopamine and these responses appeared not to be correlated with those effected by the catecholamines. A further separation of the actions of octopamine and the catecholamines was evident when the amine induced responses were compared in the presence of the antagonists, propranolol and α-flupenthixol. These blocking compounds were effective in attenuating the effects of the catecholamines, but had no effect upon the octopamine induced changes in firing rate. The results suggest that receptors sensitive to octopamine and which appear to be pharmacologically distinct from those previously categorized as catecholamine receptors, may exist on central neurones of the rat. On the basis of the present findings, it was evident that when the technique of iontophoresis is combined with standard neurophysiological methods of identifying central neurones by their responses to synaptic stimulation, valuable information can be obtained concerning the nature of the synaptic transmitters employed by these cells. / Medicine, Faculty of / Cellular and Physiological Sciences, Department of / Graduate

Myelin and oligodendrocytes (OLGs) are the apparent targets of the immune-mediated injury that underlies the development of multiple sclerosis (M8). Recovery from M8 clinical relapses likely reflects remyelination attributed to recruitment and differentiation of oligodendrocyte progenitor cells (OPCs), rather than to new process formation by previously myelinating OLGs. Newly emerging M8-directed immuno-modulatory therapies (statins and FTY720) can readily cross the blood-brain barrier and have been shown to impact signaling pathways implicated in cytoskeletal regulation, differentiation, migration, and survival; these are cellular events presumably important for myelin integrity and remyelination. / Statins inhibit the production of cholesterol (concentrated in the myelin membrane) and isoprenoids (post-translational attachments regulating the functions of proteins such as the Rho GTPases). We showed that treatment of human and rodent-derived OPCs with lipophilic statins induced an initial process extension associated with enhanced differentiation and impaired spontaneous migration, whereas prolonged treatment induced process retraction and cell death. Rodent and human mature OLGs demonstrated similar cytoskeletal and survival responses. Chronic simvastatin therapy of mice inhibited remyelination following demyelination induced by the OLG toxin, cuprizone, attributed to a block in OPC differentiation and consequent decrease in mature OLGs. Even fully myelinated animals treated with simvastatin over the long-term demonstrated a decrease in myelin in the brain by maintaining oligodendroglial cells in the pre-OLG state and preventing continual replacement of mature OLGs. / FTY720 is an agonist of G-protein-coupled receptors S1P1, 3, 4, and 5, that are associated with distinct receptor isotype-selective activation of Rho GTPases. In human OPCs, FTY720 could induce initial S1P3/5-dependent process retraction associated with an inhibition of differentiation, and subsequent S1P1-dependent process extension. Mature OLGs showed a dose-dependent cyclic modulation of process extension and retraction was observed over time. Both human OPCs and OLGs were rescued by FTY720 under death-promoting environments. Both cell types also demonstrated a cyclic and reciprocal modulation of S1P1 and S1P5 mRNA levels, reflected in the recurring receptor isotype-dependent functional responses over time. Studies using organotypic cerebellar slice cultures demonstrated that FTY720 did not impact myelin integrity under basal conditions, yet accelerated remyelination following lysolecithin-induced demyelination. Both treatment regimens were associated with an extension of OPC and mature OLG processes. / Our observations demonstrate that drug concentrations used to modulate immune function can have differential dose and time-dependent effects on OPCs, OLGs, as well as on myelin and remyelination processes. Our findings indicate the need to monitor the effects of putative immuno-modulatory therapies on myelin-related processes in MS patients.

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