NYMC Student Theses and Dissertations

Date of Award

6-2025

Document Type

Doctoral Dissertation - Restricted (NYMC/Touro only) Access

Degree Name

Doctor of Philosophy

Department

Physiology

First Advisor

Christopher Leonard, Ph.D.

Abstract

Serotonergic dorsal raphe neurons of the midbrain regulate numerous brain functions, including sleep-wake states. Activation of α1 receptors located on dorsal raphe neurons produces a noisy depolarization of serotonergic neurons and prolongs the postspike after hyperpolarization, but how these actions interact to influence the encoding of inputs into the firing of action potentials is not well understood. Prior studies in pyramidal neurons indicate that combination of synaptic noise and AHP magnitude can divisively or multiplicatively alter firing gain. Thus, in Aim 1 we investigated how α1 receptor activation alters spike encoding in serotonergic dorsal raphe neurons. Using whole cell recordings in brain slices, we found that phenylephrine (α1 receptor agonist) actions resulted in decreased steady state, but not initial, firing rate to enhance spike frequency adaptation over a wide range of input currents. Further, these actions resulted in a subtractive effect on steady state firing gain such that more current was required to drive firing. Since spike frequency adaptation functions as a high-pass filter, we suggest the collective actions of phenylephrine on dorsal raphe serotonergic neurons are to accentuate this high-pass filter to preserve or enhance encoding of rapidly varying inputs, like those related to transient behavioral events, while suppressing sensitivity to slowly varying inputs. These actions mirror those of orexin receptor activation on dorsal raphe neurons and are expected to be directly and indirectly disrupted by loss of orexin signaling. Orexin deficiency causes type 1 narcolepsy, a sleep disorder characterized by fragmented sleep-wake states, excessive daytime sleepiness, and cataplexy. Cataplexy episodes are bouts of muscle atonia without loss of consciousness and can be triggered by salient emotional states. Orexin modulation of dorsal raphe serotonergic neurons has been implicated in cataplexy, but inputs to dorsal raphe comprising the dysregulated circuit in the absence of orexin remain to be identified. Evidence exists that prefrontal cortex inhibition suppresses cataplexy and since its afferents project to dorsal raphe, in Aim 2 we investigated whether this circuit was involved in mediating cataplexy. We hypothesized that in the absence of orexinergic signaling, activation of prefrontal cortex inputs to dorsal raphe neurons would lead to greater disynaptic inhibition and promote cataplexy. To test this, we expressed light-gated excitatory channels (Chronos) in prefrontal cortex neurons and optically stimulated axon terminals in dorsal raphe in wild type and orexin peptide null mice while observing cataplexy-like episodes. To observe emotionally induced cataplexy effects, chocolate was provided at dark onset. We observed no effects of activating the prefrontal cortex to dorsal raphe circuitry on spontaneous, or emotionally induced, cataplexy bout frequency or duration. We then sought to confirm whether inhibition of prefrontal cortex would suppress cataplexy. We tested this by expressing excitatory DREADD receptors in prefrontal cortex GABA neurons so that we could suppress output while observing behavior following dark onset, with and without chocolate. We were not able to detect effects on cataplexy expression resulting from activating local GABA prefrontal cortex neurons, which contradicts the limited literature. We suggest alternate pathways which may be involved in cataplexy modulation by orexin for future study. Prefrontal cortex has been linked to mood disorders with corresponding sleep abnormalities, and is capable of regulating consciousness, but whether direct projections to dorsal raphe modulate arousal is unknown. In Aim 3 we tested this possibility by expressing Chronos in prefrontal cortex neurons and optically stimulating terminals in dorsal raphe in wild type and orexin-null mice while observing behavior after lights off. Sleep-like behavior (bout number, total duration) increased and latency to first epoch reduced in wild type mice, even when access to chocolate was provided. These effects were absent in orexin-null mice, where conversely, stimulation resulted in less time spent in sleep-like behavior and did not overcome the arousing effect of chocolate. Together the findings demonstrate prefrontal cortex output to dorsal raphe increases propensity for sleep-like episodes and this is sufficiently powerful to suppress wake-promoting effects of chocolate, and notably this is a novel circuit for arousal regulation which requires orexin neuropeptides. Combined, our results suggest that during aroused states, activation of orexin and alpha-1 receptors excite serotonergic dorsal raphe neurons and tune their temporal responsiveness by accentuating their high-pass filter characteristics to favor spiking in response to higher-frequency synaptic inputs. Prefrontal cortex circuitry to dorsal raphe did not appear to be involved in regulating cataplexy-like behavior in orexin null mice, but considerably increased sleep-like behavior in wild type mice even under conditions of high emotional arousal. This suggests the circuitry from prefrontal cortex to dorsal raphe can provide a signal that increases sleep pressure, a novel finding. Surprisingly, this signaling required orexin, which may be necessary for serotonergic dorsal raphe neurons to adequately encode sleep need.

Disciplines

Life Sciences | Medicine and Health Sciences

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