CYCLIC ADP-RIBOSE AND CALCIUM-INDUCED CALCIUM-RELEASE REGULATE NEUROTRANSMITTER RELEASE AT A CHOLINERGIC SYNAPSE OF APLYSIA

1. Presynaptic injection of cyclic ADP-ribose (cADPR), a modulator of the ryanodine receptor, increased the postsynaptic response evoked by a presynaptic spike at an identified cholinergic synapse in the buccal ganglion of Aplysia californica. 2. The statistical analysis of long duration postsynaptic responses evoked by square depolarizations of th e voltage-clamped presynaptic neurone showed that the number of evoked acetylcholine (ACh) quants released was increased following cADPR inj ection. 3. Overloading the presynaptic neurone with cADPR led to a tra nsient increase of ACh release followed by a depression. 4. cADPR inje ctions did not modify the presynaptic Ca2+ current triggering ACh rele ase. 5. Ca2+ imaging with the fluorescent dye rhod-2 showed that cADPR injection rapidly increased the free intracellular Ca2+ concentration indicating that the effects of cADPR on ACh release might be related to Ca2+ release from intracellular stores. 6. Ryanodine and 8-amino-cA DPR, a specific antagonist of cADPR, decreased ACh release. 7. ADP-rib osyl cyclase, which cyclizes NAD(+) into cADPR, was present in the pre synaptic neurone as shown by reverse transcriptase-polymerase chain re action experiments. 8. Application of NAD(+) the substrate of ADP-ribo syl cyclase, increased ACh release and this effect was prevented by bo th ryanodine and 8-amino-cADPR. 9. These results support the view that Ca2+-induced Ca2+ release might be involved in the build-up of the Ca 2+ concentration which triggers ACh release, and thus that cADPR might have a role in transmitter release modulation.