PDF receptor-expressing neurons in the ellipsoid body: a link between the locomotor and sleep circuits?

PÍREZ, N.; GRIFFITH, L.C.

Cold Spring Harbor, New York, Estados Unidos

Conferencia; Neurobiology of Droshophila Meeting; 2011

Cold Spring Harbor Laboratory

It has been recently been shown that some clock cells expressing the GABAA receptor subunit encoded by the Rdl gene also express the PDF receptor. Additionally, pdfr-GAL4 drives expression in cells that innervate the ellipsoid body, a component of the central complex. This, as well as the suggestion that this structure is a target of circadian clock neurons, suggest an important link between the locomotor and sleep circuit. The primary goal of these experiments is to study this connection. In addition, we are interested in analyzing the effect that sleep deprivation has on the activity of this circuit. To approach these issues, we performed whole-brain optical imaging experiments using flies expressing the FRET-based cAMP-sensitive sensor Epac1-camps under the control of the PDF receptor promoter. In previous unpublished experiments we have found that bath application of the PDF peptide modulates the response of these cells to other bioamines, but did not cause a direct change in calcium levels. In the current experiments, male fly brains were dissected and pinned to a chamber that was constantly perfused with artificial hemolymph. Using a cAMP reporter we show that bath application of PDF (10 M) causes a significant increase in the cAMP level of these cells measured as the ratio of YFP/CFP. When 1 µM TTX was included in the bath to block voltage-gated Na+ channels, we found that response elicited by PDF was not eliminated. We also tested the effects of other neurotransmitters on ellipsoid body neurons. For example, 20 mM acetylcholine (ACh) caused a significant increase in cAMP levels, and the temporal dynamics of this response were affected by time of day. In order to test the effect that sleep deprivation had on the physiology of these cells, we performed behavioral experiments in which flies were sleep deprived for twelve hours prior to imaging. The imaging sessions were performed either immediately after the end of deprivation period or following a 4 hour recovery period. Although the effect of sleep deprivation is clear at the behavioral level, the physiological responses of these cells to ACh remained similar to non-sleep deprived controls. More experiments are being conducted to elucidate the mechanism of action of these cells, as well as to test the effects that other neurotransmitters, such as the wake-promoting bioamine dopamine, have in this cell population. Additionally, experiments using PDF receptor mutants are being performed to test the effect that removing the receptor has on the observed responses.