As the organ system responsible for controlling our movements, senses, and consciousness, the nervous system must process and transmit information rapidly. To meet this demand, nervous systems utilize networks of nerve cells, each capable of generating electrical impulses and transmitting them to other cells in the network. Understanding the mechanisms underlying the generation and transmission of these impulses is thus essential to understanding fundamental mechanisms of neural function. The focus of our laboratory has been on the mechanisms of signal transmission, specifically those occuring at chemical synapses. Here chemical neurotransmitter is released from the presynaptic neuron and activates the postsynaptic cell membrane. One interesting aspect of this process is the highly regulated and rapid form of exocytosis responsible for neurotransmitter release.

Although the molecular mechanisms underlying neurotransmitter release remain incompletely understood, recent progress has implicated a number of identified proteins. Ideally, the functions of these and other proteins in the release process can be defined by specific perturbation of individual gene products, followed by functional analysis at native synapses in vivo. Our laboratory utilizes the fruit fly, Drosophila melanogaster, as a model experimental system in which synaptic mechanisms similar to those of vertebrates can be studied in vivo, using a powerful combination of genetic, molecular, biochemical, electrophysiological and ultrastructural approaches. Genetic methods are used to screen for mutants defective in synaptic transmission; molecular and biochemical methods are used to identify, characterize, and manipulate the affected protein; and electrophysiological and ultrastructural methods are used to investigate the in vivo function of the protein at native synapses.

Ongoing projects include the analysis of temperature-sensitive (TS) paralytic mutants exhibiting conditional defects in synaptic transmission. These conditional mutants allow normal development and function at permissive temperature, while also allowing acute perturbation of a specific gene product in the mature animal. These features make TS paralytic mutants a unique and powerful tool for analyzing the in vivo physiological functions of specific proteins. One example is our analysis in comatose mutants (see Figure), in which paralysis results from a TS defect in a Drosophila N-ethylmaleimide sensitive fusion protein (NSF). Our functional analysis, together with biochemical analysis in comatose [Tolar and Pallanck (1998) Journal of Neuroscience 18(24):10250-10256] have defined the function of this NSF protein in regulated exocytosis of neurotransmitter. Subsequently we have extended our analysis by analyzing TS mutations affecting other key components of the neurotransmitter release apparatus. For example we have isolated a TS allele of the cacophony gene, which encodes the primary structural subunit of a voltage-gated calcium channel, and have utilized this mutant to demonstrate that cacophony encodes a primary synaptic calcium channel functioning in neurotransmitter release. Further detailed analysis of TS mutations alone and in combination promises to provide new insights into the in vivo functions and interactions of specific gene products in synaptic transmission.