Abstract
Prejunctional nicotinic acetylcholine receptors (nAChRs) amplify postganglionic sympathetic neurotransmission, and there are indications that intraterminal Ca2+ stores might be involved. However, the mechanisms by which nAChR activation stimulates neurotransmitter release at such junctions is unknown. Rapid local delivery (picospritzing) of the nAChR agonist epibatidine was combined with intracellular sharp microelectrode recording to monitor spontaneous and field-stimulation-evoked neurotransmitter release from sympathetic nerve terminals in the mouse isolated vas deferens. Locally applied epibatidine (1 µM) produced ‘epibatidine-induced depolarisations’ (EIDs) that were similar in shape to spontaneous excitatory junction potentials (SEJPs) and were abolished by nonselective nAChR antagonists and the purinergic desensitizing agonist α,β-methylene ATP. The amplitude distribution of EIDs was only slightly shifted towards lower amplitudes by the selective α7 nAChR antagonists α-bungarotoxin and methyllcaconitine, the voltage-gated Na+ channel blocker tetrodotoxin or by blocking voltage-gated Ca2+ channels with Cd2+. Lowering the extracellular Ca2+ concentration reduced the frequency of EIDs by 69%, but more surprisingly, the Ca2+-induced Ca2+ release blocker ryanodine greatly decreased the amplitude (by 41%) and the frequency of EIDs by 36%. Ryanodine had no effect on electrically-evoked neurotransmitter release, paired-pulse facilitation, SEJP frequency, SEJP amplitude or SEJP amplitude distribution. These results show that activation of non-α7 nAChRs on sympathetic postganglionic nerve terminals induces high-amplitude junctional potentials that are argued to represent multipacketed neurotransmitter release synchronized by intraterminal Ca2+-induced Ca2+ release, triggered by Ca2+ influx directly through the nAChR. This nAChR-induced neurotransmitter release can be targeted pharmacologically without affecting spontaneous or electrically-evoked neurotransmitter release.
Highlights
Activation of nicotinic acetylcholine receptors (nAChRs) located on nerve terminals is an important mechanism that modulates neurotransmitter release
There was no significant difference between spontaneous excitatory junction potentials (SEJPs) amplitude distributions recorded in the presence and absence of ryanodine (P = 0.12; Fig. 4E). These results indicate that epibatidine-induced depolarisations (EIDs) are partially dependent on Ca2+-induced Ca2+ release (CICR), whereas electrically-evoked neurotransmitter release and spontaneous neurotransmitters release do not involve CICR
This study has shown that rapid application of epibatidine caused a series of smooth muscle cell depolarisations similar in appearance to SEJPS that were sensitive to nonselective nAChR antagonists and blocked following desensitisation of P2X1 receptors
Summary
Activation of nAChRs located on nerve terminals is an important mechanism that modulates neurotransmitter release. Most research on the mechanism of nAChR-induced modulation of neurotransmitter release has been carried out in the CNS, where a number of mechanisms have been proposed including initiation of action potentials [5], activation of voltage-gated Ca2+ channels [VGCCs; 6], direct influx of Ca2+ through the nAChR [7], mobilisation of intraneuronal Ca2+ stores [8], an unidentified NO-dependent process [9] and a number of Ca2+-dependent cellular processes, such as recruitment of protein kinases [see 10]. Previous studies in the rodent vas deferens have shown that activation of nAChRs can both induce neurotransmitter release and potentiate electrically-evoked neurotransmitter release [12,13,14,15], the precise mechanism of the modulation remains unknown
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