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Crustacean muscles are innervated by phasic and tonic motor neurons that display differential physiology and have morphologically distinct synaptic terminals. Phasic motor neurons release much more transmitter per impulse and have filiform terminals, whereas tonic motor neurons release less transmitter and have larger terminals with prominent varicosities. Using an antibody raised against Drosophila frequenin (frq), a calcium-binding protein that enhances transmitter release in Drosophila synaptic terminals, we found that frq-like immunoreactivity is prominent in many of the phasic, but not tonic nerve endings of crayfish motor neurons. In contrast, synapsin- and dynamin-like immunoreactivities are strongly expressed in both types of terminal. The immunocytochemical findings strongly suggested the presence of an frq-like molecule in crayfish, and its differential expression indicated a possible modulatory role in transmitter release. Therefore, we cloned the cDNA sequences for the crayfish and lobster homologues of Drosophila frq. Crustacean frequenins are very similar in sequence to their Drosophila counterpart, and calcium-binding regions (EF hands) are conserved. The widespread occurrence of frq-like molecules and their differential localization in crayfish motor neurons indicate a significant role in physiology or development of these neurons.

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We investigated synaptic ultrastructure of individual nerve ending varicosities at the Drosophila larval neuromuscular junction in transgenic larvae overexpressing the learning gene dunce (dnc) in the nervous system. It was previously shown that cAMP is reduced to one‐third normal in these larvae and that they have fewer nerve terminal varicosities and smaller junction potentials, although transmitter release from individual nerve ending varicosities is not significantly altered. We tested the hypothesis that synaptic ultrastructure is modified to compensate for possible reduced efficacy of synaptic transmission resulting from lower than normal cAMP. Synaptic size and number of presynaptic dense bodies (active zone structures) per synapse are modestly enhanced in transgenic larvae overexpressing the dnc gene product and in rutabaga (rut1 ) mutant larvae, which have reduced adenylyl cyclase activity and reduced neural cAMP. The incidence of complex synapses (possessing 2 or more presynaptic dense bodies) was not consistently different in experimental larvae compared to controls. The observations suggest that chronic reduction of cAMP levels in the nervous system of Drosophila larvae, although leading to a modest compensatory change in synaptic structure, does not markedly alter several synaptic ultrastructural parameters which are thought to influence the strength of transmitter release; thus, homeostatic mechanisms do not act to maintain normal‐sized junction potentials by altering synaptic structure. © 2000 John Wiley & Sons, Inc. J Neurobiol 43: 89–97, 2000

The sluggish-A (slgA) gene of Drosophila melanogaster has been shown to encode for the enzyme proline oxidase, a mitochondrial enzyme which catalyzes the first step in the conversion of L-proline to L-glutamate. The slgA transcript is expressed in both larval and adult Drosophila melanogaster. Mutations in this gene lead to reduced proline oxidase activity and an elevation of free proline levels. Adult mutant flies show a striking reduction of motor activity. Since proline oxidase may contribute to the supply of the neurotransmitter glutamate in the nervous system, a reduction in proline oxidase activity could reduce neural glutamate pools and affect synaptic transmission in neurons utilizing glutamate as a transmitter, including peripheral motor neurons. We tested the hypothesis that glutamate, and synaptic transmission mediated by glutamate, are reduced at synapses of glutamatergic motor neurons in slgA mutants. Levels of glutamate and proline in different cell compartments, and functional properties of synaptic transmission were compared in slgA and control specimens. Proline is elevated in muscle cells of slgA mutants, indicating that the slgA gene regulates tissue proline levels. In nerve terminal varicosities, proline levels were low in both mutants and controls. Glutamate levels in nerve terminal varicosities of slgA mutants and controls were similar. In addition, we found that glutamatergic synaptic transmission at individual nerve endings and at the whole-cell level was similar in slgA mutants and controls. Thus, proline oxidase does not play a major role in generating neuronal glutamate pools at the Drosophila larval neuromuscular junction, and larval neuromuscular performance is not altered significantly in slgA mutants. Metabolic pathways other than that involving proline oxidase are able to sustain glutamatergic synaptic function in Drosophila larvae.