Effects of gap junction misexpression on synapses between auditory sensory neurons and the giant fiber of Drosophila melanogaster

The synapse between auditory Johnston’s Organ neurons (JONs) and the giant fiber (GF) of Drosophila is structurally mixed, being composed of cholinergic chemical synapses and Neurobiotin-(NB) permeable gap junctions, which consist of the innexin Shaking-B (ShakB). Misexpression of one ShakB isoform, ShakB(N+16), in a subset of JONs that do not normally form gap junctions, results in their de novo dye coupling to the GF. This is similar to the effect of misexpression of the transcription factor Engrailed (En) in these same neurons, which also causes the formation of additional chemical synapses. In order to test the hypothesis that ShakB misexpression would similarly affect the distribution of chemical synapses, fluorescently-labeled presynaptic active zone protein (Brp) was expressed in JONs and the changes in its distribution were assayed with confocal microscopy. Both ShakB(N+16) and En increased the dye-coupling of JONs with the GF, indicating the formation of ectopic gap junctions. Conversely, expression of the ‘incorrect’ isoform, ShakB(N) abolishes dye coupling. However, while En misexpression increased the chemical contacts with the GF and the amount of GF medial branching, ShakB misexpression did not. ShakB immunocytochemistry showed that misexpression of ShakB(N+16) increases gap junctional plaques in JON axons but ShakB(N) does not. We conclude that both subsets of JON form chemical synapses onto the GF dendrites but only one population forms gap junctions, comprised of ShakB(N+16). Misexpression of this isoform in all JONs does not result in the formation of new mixed synapses but in the insertion of gap junctions, presumably at the sites of existing chemical synaptic contacts with the GF.

Opioids potentiate electrical transmission at mixed synapses on the Mauthner cell

Opioid receptors were shown to modulate a variety of cellular processes in the vertebrate central nervous system, including synaptic transmission. While the effects of opioid receptors on chemically mediated transmission have been extensively investigated, little is known of their actions on gap junction-mediated electrical synapses. Here we report that pharmacological activation of mu-opioid receptors led to a long-term enhancement of electrical (and glutamatergic) transmission at identifiable mixed synapses on the goldfish Mauthner cells. The effect also required activation of both dopamine D1/5 receptors and postsynaptic cAMP-dependent protein kinase A, suggesting that opioid-evoked actions are mediated indirectly via the release of dopamine from varicosities known to be located in the vicinity of the synaptic contacts. Moreover, inhibitory inputs situated in the immediate vicinity of these excitatory synapses on the lateral dendrite of the Mauthner cell were not affected by activation of mu-opioid receptors, indicating that their actions are restricted to electrical and glutamatergic transmissions co-existing at mixed contacts. Thus, as their chemical counterparts, electrical synapses can be a target for the modulatory actions of the opioid system. Because gap junctions at these mixed synapses are formed by fish homologs of the neuronal connexin 36, which is widespread in mammalian brain, it is likely that this regulatory property applies to electrical synapses elsewhere as well.

Mixed Synapses as a Testbed for Developing Reagents for Building Connectomes and Synaptomes

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.