Light buffers the wake‐promoting effect of dopamine

Sleep is driven and regulated by the integration of diverse internal and external (environmental) cues. Light is known to be a potent inhibitor of sleep in diurnal animals (awake during daylight hours and sleep at night), including both humans and fruit flies. Yet wakefulness does not scale linearly with light intensity and a lack of light does not automatically result in sleep. (Evolution seems unlikely to favor animals who become hyperactive in dangerously hot midday sunlight and fall asleep in an uncontrollable narcoleptic fashion when the sun goes down, unable to wake until the next morning.) The sleep regulatory system must be plastic — capable of weighing the relative importance of incoming sleep and wake‐promoting cues, and buffering the effects of those cues on sleep drive accordingly. In a recent Nature Neuroscience paper from a team led by postdoc Yuhua Shang (Rosbash lab), with collaborators from the Griffth, Pollack, and Hong labs at Brandeis, we determined at the cell and molecular level how the fruit fly, Drosophila melanogaster, is able to buffer the wake‐promoting effects of the neurotransmitters dopamine and octopamine in the presence of light in order to maintain a proper sleep:wake balance.

It is known that dopamine and octopamine both promote wakefulness in flies. Previous work in the Rosbash and Griffith labs has shown that 10 neurons in the Drosophila brain that release the neuropeptide pigment‐dispersing factor (PDF), known as the l‐LNvs, are critical for transducing the wake‐promoting effects of light. Quantifying mRNAs from all 18 PDF-expressing neurons revealed an enrichment of octopamine and dopamine receptors specifically in the ten wake‐promoting l‐LNvs. We wondered if the l‐LNvs were also able to respond to and transduce the wake‐promoting effects of dopamine and octopamine, and if so, how these effects were integrated with the wake‐promoting effects of light by these cells.

Figure: The l-LNvs use two parallel intracellular pathways to regulate the stimulating effects of DA and OA. Both DA and OA increase the cAMP levels in the l-LNvs. Light in the housing environment suppresses the effects of both DA and OA, but in different ways. In the case of dopamine, light induces increased expression of an inhibitory D2R receptor and in the case of octopamine, the effect is dependent on the circadian clock (Per.)

Using a fluorescence resonance energy transfer (FRET)‐based cyclic AMP reporter expressed in all 18 Pdf neurons, we were able to see robust responses to both octopamine and dopamine in only the t0 l‐LNvs, confirming the mRNA result. To verify that the l‐LNvs are in fact in close apposition to presynaptic octopaminergic and dopaminergic neurons, we looked for reconstitution of a split GFP protein between pre- and post‐synaptic cells. With different GFP fragments expressed at the membrane of the l‐LNvs and presynaptic dopaminergic or octopaminergic neurons, reconstituted GFP would only be visible if these cell populations were in close contact. Reconstituted GFP was seen in both cases around l‐LNv cell bodies and dendritic areas.

To determine the behavioral effect of increased dopaminergic neuron activity on sleep, we transiently hyper‐excited the dopaminergic neurons in flies using the Garrity lab’s heat‐activated dTrpA1 channel. When the housing temperature of flies expressing dTrpA1 in dopaminergic neurons was increased, activating dTrpA1 activity, flies exhibited increased wakefulness. Interestingly, this increased wakefulness was much greater in flies housed in constant darkness as compared to those housed in light:dark cycling conditions. This suggested that the l‐LNvs are a convergence point for the wakepromoting effects of dopamine and light. FRET analysis confirmed this, showing that the l‐LNv response to both dopamine and octopamine is much weaker in flies kept in light:dark conditions as compared to those kept in constant darkness. We then determined that light causes increased expression of an inhibitory dopamine receptor, resulting in a weaker excitatory response to dopamine by the l‐LNvs. In the case of octopamine, the circadian clock was found to regulate the effects of light. Such plasticity allows flies to maintain similar amounts of total sleep in varying environmental conditions, decreasing the relevance of internally generated wake‐promoting cues, in the presence of stronger environmental cues (light). It will be interesting to see how these results generalize to mammals, since light and dopamine also both promote wakefulness in mammals.

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