Single CNS neurons can be dissociated with preserved adherent functional synaptic boutons without using any enzyme, namely when preparing a "synaptic bouton" preparation. This allows experimenters to investigate the effects of presynaptic modulators of synaptic transmission with unprecedented case and accuracy. Moreover, a single bouton can be visualizecd using fluorescent markers and can also be focally stimulated with electrical pulses. In this communication, high voltage-dependent <$E roman Ca sup 2+> channels of nerve endings, as one of the experimental examples using the "synaptic bouton" preparation, are described. Calcium channels belonging to different subtypes, which trigger GABA release from nerve terminals (boutons) projecting to rat hippocampal CA1 pyramidal neurons, were studied. GABA-ergic evoked inhibitory postsynaptic currents (eIPSCs) were recorded; these currents were elicited by focal stimulation of single boutons in mechanically dissociated neurons and by stimulation of a nerve bundle in slice preparations. Nilvadipine, an L-type <$E roman Ca sup 2+> channel blocker, completely inhibited eIPSCs evoked by stimulation of single boutons but exerted no effect on eIPSCs evoked by low-frequency stimulation of the nerve bundle. Nilvadipine did, however, prevent potentiation of the eIPSC amplitude following high-frequency stimulation of the nerve bundles in slice preparations. <$E omega>-Conotoxin-GVIA, a blocker of N-type <$E roman Ca sup 2+> channels, and <$E omega>-agatoxin-IVA, a blocker of P/Q-type <$E roman Ca sup 2+> channels, completely inhibited eIPSCs in 33,3 and 83,3 % of the recordings from single bout

ons, respectively. In response to low-frequency nerve bundle stimulation in the slice preparation, both <$E omega>-conotoxin-GVIA and <$E omega>-agatoxin-IVA partially reduced the amplitude of eIPSCs, and the residual component could be abolished by <$E roman Cd sup 2+>. From these results, the following hypotheses could be drawn, (i) The distribution of P/Q- and N-type <$E roman Ca sup 2+> channels at a single bouton is nonuniform; (ii) when a focal stimulation is applied to a single bouton, L-type <$E roman Ca sup 2+> channels play a significant role in generation of action potentials, which subsequently activate P/Q- and N-type <$E roman Ca sup 2+> channels at GABA release sites; and (iii) action potentials conducted through axons in the slice preparation are sufficient to depolarize the bouton membrane, even when L-type <$E roman Ca sup 2+> channels are suppressed.

Ключ. слова: mechanical dissociation of CNS neurons, functional synaptic bouton, focal stimulation, Ca"2+ channel subtypes
Індекс рубрикатора НБУВ: Е60*739.12

Рубрики:

Загальна зоологія

Шифр НБУВ: Ж61875 Пошук видання у каталогах НБУВ 

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