Glutamate, previously demonstrated to participate in regulation of the resting membrane potential in skeletal muscles, also regulates non-quantal acetylcholine (ACh) secretion from rat motor nerve endings. Non-quantal ACh secretion was estimated by the amplitude of endplate hyperpolarization (H-effect) following blockade of skeletal muscle post-synaptic nicotinic receptors by (+)-tubocurarine and cholinesterase by armin (diethoxy-p-nitrophenyl phosphate). Glutamate was shown to inhibit non-quantal release but not spontaneous and evoked quantal secretion of ACh. Glutamate-induced decrease of the H-effect was enhanced by glycine. Glycine alone also lowered the H-effect, probably due to potentiation of the effect of endogenous glutamate present in the synaptic cleft. Inhibition of N-methyl-D-aspartate (NMDA) receptors with (+)-5-methyl-10,11-dihydro-5H-dibenzocyclohepten-5,10-imine (MK801), DL-2-amino-5-phosphopentanoic acid (AP5) and 7-chlorokynurenic acid or the elimination of Ca It is well known that glutamate serves as a neurotransmitter at invertebrate neuromuscular junctions (Lunt and Olsen 1988) whereas acetylcholine (ACh) serves that role in the vertebrate. However, investigations during the last decade indicate a role for glutamate as a signalling molecule at the neuromuscular junction of vertebrates (see for review Grozdanovic and Baumgarten 1999). In support of this notion, glutamate transporter mRNA has been shown to be present in the cytoplasm of rat spinal motoneurones along with a glutamate-like immunoreactivity (Meister et al. 1993). Glutamate has been found in nerve terminals of rat motoneurones in association with synaptic vesicles (Waerhaug and Ottersen 1993) and evidence that it is co-released with ACh in cholinergic nerve terminals has been documented (Vyas and Bradford 1987;Israel et al. 1993;Meister et al. 1993). In addition, N-methyl-D-aspartate (NMDA) and Received August 15, 2002; revised manuscript received November 20, 2002; accepted December 19, 2002. Address correspondence and reprint requests to Dr Albert K. Urazaev, Department of Physiology, The Brody School of Medicine, East Carolina University, Greenville, NC 27858, USA. E-mail: urazaeva@mail.ecu.eduAbbreviations used: ACh, acetylcholine; AP5, DL-2-amino-5-phosphopentanoic acid; D-NAME, N G -nitro-D-arginine methyl ester; EPP, endplate potentials; L-NAME, N G -nitro-L-arginine methyl ester; mEPP, miniature endplate potentials; MK801, (+)-5-methyl-10,11-dihydro-5H-dibenzocyclohepten-5,10-imine; NO, nitric oxide; ODQ, 1H [1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one.

Noradrenaline (NA) increases synaptic efficacy at the frog neuromuscular junction. To test the hypothesis that one of the actions of NA is to shorten the period over which evoked quanta are released, we measured the latencies of focally recorded uniquantal endplate currents (EPCs). NA shortened the release period for evoked quantal release. The interval between the time when responses with minimal delay appeared and the point at which 90 % of all latencies had occurred was shortened in the presence of 1 × 10−5 M NA by about 35 % at 20 °C and by about 45 % at 8 °C. Inhibitor and agonist experiments showed that NA acts on a β‐adrenoreceptor. The better synchronization of release significantly increased the size of reconstructed multi‐ quantal EPCs. This suggests that NA facilitates synaptic transmission by making the release of quanta more synchronous. The synchronizing action of NA might potentiate neuromuscular transmission during nerve regeneration, transmitter exhaustion and other extreme physiological states where the quantal content is reduced, such as survival in cold and hibernation.

After anticholinesterase treatment, the postsynaptic muscle membrane is depolarized by about 5 mV due to nonquantal release of acetylcholine (ACh) from the motor nerve terminal. This can be demonstrated by the hyperpolarization produced by the addition of curare (H-effect). The magnitude of the H-effect was decreased significantly to 3 mV when the nitric oxide (NO) donors, sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine (SNAP) were applied to the muscle, or when NO production was elevated by adding L-arginine, but not D-arginine, as a substrate. The H-effect was increased to 8-9 mV by inhibition of NO synthase by L-nitroarginine methylester (L-NAME), or by guanylyl cyclase inhibition by methylene blue and 1H-[1,2,4]oxidiazolo[4,3-a]quinoxalin-1-one (ODQ). ODQ increased the H-effect to 7.3 +/- 0.2 mV and diminished the SNP-induced decrease of the H-effect when applied together with SNP. The effects of NO donors and L-arginine were eliminated by adding reduced haemoglobin, an extracellular NO scavenger. The present results, together with earlier evidence for the presence of NO synthase in muscle fibres, indicate that nonquantal release of ACh is modulated by NO production in the postsynaptic cell.