Reciprocal Lateral Hypothalamic and Raphe GABAergic Projections Promote Wakefulness

Abstract

The lateral hypothalamus (LH), together with multiple neuromodulatory systems of the brain, such as the dorsal raphe nucleus (DR), is implicated in arousal, yet interactions between these systems are just beginning to be explored. Using a combination of viral tracing, circuit mapping, electrophysiological recordings from identified neurons, and combinatorial optogenetics in mice, we show that GABAergic neurons in the LH selectively inhibit GABAergic neurons in the DR, resulting in increased firing of a substantial fraction of its neurons that ultimately promotes arousal. These DR GABA neurons are wake active and project to multiple brain areas involved in the control of arousal, including the LH, where their specific activation potently influences local network activity leading to arousal from sleep. Our results show how mutual inhibitory projections between the LH and the DR promote wakefulness and suggest a complex arousal control by intimate interactions between long-range connections and local circuit dynamics.

Activity of the Lateral Hypothalamus during Genetically Determined Absence Seizures

“…We used viral tracing and ChR2-assisted circuit mapping to show that a population of LH neurons establish monosynaptic connections with DRN 5-HT neurons. While previous studies showed that LH GABAergic neurons preferentially contact DRN GABAergic neurons 23 here we present evidence for a direct excitatory LH to DRN connection mediated through iGluR receptors directed towards 5-HT neurons. The exact neurochemical identity of this subset of DR-projecting LH neurons remains to be established given the numerous subpopulations among hypothalamic neurons [33].…”

“…Injection of AAV-syn-ChR2-Venus [29] into the LH resulted in somatic ChR2-Venus expression in the LH region and dense axonal expression in the brainstem area including the DRN [23]. To confirm the results of the anterograde tracing we injected the retrograde viral vector PRV-cre [30] in the DRN area (Figure 1A) of tdTomato reporter mice (Ai9, RCL-tdTomato, Jackson Laboratory, Bar Harbor, ME, USA).…”

“…For assessing the effect of local photostimulation of ChR2 expressing LH axons in the DRN we used methods similar to those previously described [23]. Briefly, male WT mice were injected bilaterally with 50 nl of AAV-syn-ChR2-Venus [29] in the LH at the following coordinates (AP: −1.4, ML: 1, DV: −5.2 all mm from Bregma) using a nanoinjector (Nanoliter 2000, WPI, Sarasota, FL, USA) connected to a glass pipette (~20 µm tip diameter) at a rate of 1 nl/s.…”

“…We next used optogenetics to functionally characterize the LH to DRN circuit (Figure 2A). We found that photostimulation of AAV-syn-ChR2-Venus-expressing LH terminals in the DRN evoked excitatory postsynaptic potentials (EPSPs) of 7.58 ± 0.9 mV peak amplitude with a 11.07 ± 2.46 ms decay time in 10 out of 51 neurons tested (20%) DRN cells (Figure 2D,E) and inhibitory postsynaptic potentials (IPSPs) in seven neurons (14%) [23]. Of the 10 neurons showing EPSPs five were confirmed to be DRN serotonergic neurons using post-hoc immunohistochemistry (Figure 2B,E).…”

“…One of the main inputs to the dorsal raphe nucleus (DRN) is the lateral hypothalamus (LH) [18,19], a brain area consisting of neurochemically heterogeneous neuronal populations implicated in the modulation of arousal [20] and control of energy intake [21]. GABAergic neurons in the LH selectively inhibit thalamic reticular nucleus neurons [22] and DRN GABAergic neurons [23] promoting wakefulness. The LH receives prominent cortical projections from the medial prefrontal and orbitofrontal cortices [24][25][26].…”