Electrophysiological Analysis of Synaptic Transmission in Central Neurons ofDrosophilaLarvae

Abstract

We report functional neuronal and synaptic transmission properties in Drosophila CNS neurons. Whole cell current- and voltage-clamp recordings were made from dorsally positioned neurons in the larval ventral nerve cord. Comparison of neuronal Green Fluorescent Protein markers and intracellular dye labeling revealed that recorded cells consisted primarily of identified motor neurons. Neurons had resting potentials of -50 to -60 mV and fired repetitive action potentials (APs) in response to depolarizing current injection. Acetylcholine application elicited large excitatory responses and AP bursts that were reversibly blocked by the nicotinic receptor antagonist D-tubocurarine (dtC). GABA and glutamate application elicited similar inhibitory responses that reversed near normal resting potential and were reversibly blocked by the chloride channel blocker picrotoxin. Multiple types of endogenous synaptically driven activity were present in most neurons, including fast spontaneous synaptic events resembling unitary excitatory postsynaptic currents (EPSCs) and sustained excitatory currents and potentials. Sustained forms of endogenous activity ranged in amplitude from smaller subthreshold "intermediate" sustained events to large "rhythmic" events that supported bursts of APs. Electrical stimulation of peripheral nerves or focal stimulation of the neuropil evoked sustained responses and fast EPSCs similar to endogenous events. Endogenous activity and evoked responses required external Ca(2+) and were reversibly blocked by dtC application, indicating that cholinergic synaptic transmission directly underlies observed activity. Synaptic current amplitude and frequency were reduced in shibire conditional dynamin mutants and increased in dunce cAMP phosphodiesterase mutants. These results complement and advance those of recent functional studies in Drosophila embryonic neurons and demonstrate the feasibility of in-depth synaptic transmission and plasticity studies in the Drosophila CNS.

Astrocytic glutamate transport regulates a Drosophila CNS synapse that lacks astrocyte ensheathment

“…Note that glutamate signaling in the fly CNS differs from vertebrates due to the presence of inhibitory glutamate receptors. Previous studies have shown that motor neurons express glutamate-gated chloride channels (Rohrbough and Broadie 2002), which have been characterized in other invertebrates such as C. elegans (Vassilatis et al, 1997) but are absent from vertebrate genomes (Wolstenholme, 2012). The Drosophila homolog of this receptor, GluCl-alpha, produces a glutamate-gated chloride current when cloned and expressed in Xenopus oocytes (Cully et al, 1996).…”

DrosophilaGlial Glutamate Transporter Eaat1 Is Regulated by Fringe-Mediated Notch Signaling and Is Essential for Larval Locomotion

“…These motor neurons exhibit large excitatory synaptic currents, termed SRCs (Rohrbough and Broadie, 2002), between 10 and 15 times every minute (Fig. 5 A, B) (Baines et al, 2002).…”

Cholinergic Synaptic Transmission in AdultDrosophilaKenyon CellsIn Situ

“…Behavioral studies in the fruit fly Drosophila melanogaster, using an extensive library of mutant and transgenic animals, have resulted in identification of a number of genes and signal cascades that contribute to memory formation (Davis, 2004). In addition, physiological recordings in Drosophila, at the neuromuscular junction, from neurons in primary culture and in the CNS of embryos and larvae, have expanded our understanding of the molecular mechanisms regulating synaptic plasticity and neuronal excitability (Zhong and Wu, 1991;Broadie et al, 1997;Lee and O'Dowd, 2000;Renger et al, 2000;Yao and Wu, 2001;Hodges et al, 2002;Park et al, 2002;Rohrbough and Broadie, 2002;Baines, 2003;Hou et al, 2003;Rohrbough et al, 2003). However, limited electrophysiological access to neurons in central circuits in the adult fly has precluded exploration of the direct cellular links that mediate the translation of changes in gene expression to memory formation.…”