Physiologic and nanoscale distinctions define glutamatergic synapses in tonic vs phasic neurons

Neurons exhibit a striking degree of functional diversity, each one tuned to the needs of the circuitry in which it is embedded. A fundamental functional dichotomy occurs in activity patterns, with some neurons firing at a relatively constant “tonic” rate, while others fire in bursts - a “phasic” pattern. Synapses formed by tonic vs phasic neurons are also functionally differentiated, yet the bases of their distinctive properties remain enigmatic. A major challenge towards illuminating the synaptic differences between tonic and phasic neurons is the difficulty in isolating their physiological properties. At the Drosophila neuromuscular junction (NMJ), most muscle fibers are co-innervated by two motor neurons, the tonic “MN-Ib” and phasic “MN-Is”. Here, we employed selective expression of a newly developed botulinum neurotoxin (BoNT-C) transgene to silence tonic or phasic motor neurons. This approach revealed major differences in their neurotransmitter release properties, including probability, short-term plasticity, and vesicle pools. Furthermore, Ca2+ imaging demonstrated ~two-fold greater Ca2+ influx at phasic neuron release sites relative to tonic, along with enhanced synaptic vesicle coupling. Finally, confocal and super resolution imaging revealed that phasic neuron release sites are organized in a more compact arrangement, with enhanced stoichiometry of voltage-gated Ca2+ channels relative to other active zone scaffolds. These data suggest that distinctions in active zone nano-architecture and Ca2+ influx collaborate to differentially tune glutamate release at synapses of tonic vs phasic neuronal subtypes.