Group III Metabotropic Glutamate Receptors Maintain Tonic Inhibition of Excitatory Synaptic Input to Hypocretin/Orexin Neurons

Leptin Acts via Lateral Hypothalamic Area Neurotensin Neurons to Inhibit Orexin Neurons by Multiple GABA-Independent Mechanisms

“…AAV-Flex-hM3Dq-mCherry virus (Alexander et al, 2009) was purchased from the UNC Vector Core (University of North Carolina, Chapel Hill, NC). Virus was stereotaxically injected bilaterally into the LHA of Nts cre ; OX-EGFP mice at 5-6 weeks of age.…”

“…To examine the functional connectivity between LHA Nts neurons and OX neurons, we used a recombinant adeno-associated virus (AAV) that mediates the cre-dependent expression of DREADDs (designer receptors exclusively activated by designer drugs; expressed as DREADD-mCherry fusion proteins), which are genetically engineered muscarinic receptor variants that are insensitive to endogenous ligands but are activated by the otherwise biologically inert agonist CNO. CNO activates neurons containing the G q -coupled DREADD, hM3Dq (Armbruster et al, 2007;Alexander et al, 2009;Krashes et al, 2011).…”

“…Excitatory input to OX neurons is substantially greater than inhibitory input to OX cells, and the excitatory input is modulated by a variety of stimuli (Li et al, 2002;Acuna-Goycolea and van den Pol, 2004;Acuna-Goycolea et al, 2004;Fu et al, 2004;Horvath and Gao, 2005;Rao et al, 2007Rao et al, , 2013, including short-term (24 h) fasting (Horvath and Gao, 2005), which decreases circulating leptin concentrations. To investigate whether leptin modulates excitatory input to OX neurons, we recorded sEPSCs in OX-EGFP neurons, voltage clamped to Ϫ50 mV, before and after the application of leptin (10 nM).…”

Glucagon-Like Peptide 1 Excites Hypocretin/Orexin Neurons by Direct and Indirect Mechanisms: Implications for Viscera-Mediated Arousal

“…To identify hypocretin cells, hypothalamic slices were prepared from transgenic mice [from Dr. T. Sakurai (University of Tsukuba, Tsukuba, Japan)] that selectively express GFP in hypocretin neurons (Li et al, 2002;Yamanaka et al, 2003). MCH-containing neurons were identified by selective GFP expression after recombinant adeno-associated virus (AAV-MCH) infection in vivo or by using transgenic mice that selectively express GFP in MCH cells (van den Pol et al, 2004); the electrophysiological properties of MCH neurons were similar with each method of identification. We used 2-to 3-week-old animals maintained in a normal 12 hr light/dark cycle.…”

“…All of the slice preparation procedures were performed as previously described (Li et al, 2002;Acuna-Goycolea et al, 2004). Briefly, animals were deeply anesthetized (sodium pentobarbital, 100 mg/kg), and then the brain was rapidly removed and placed in icecold oxygenated (95% O 2 -5% CO 2 ) solution that contained (in mM): 220 sucrose, 2.5 KCl, 6 MgCl 2 , 1 CaCl 2 , 1.25 NaH 2 PO 4 , 26 NaHCO 3 , and 10 glucose, pH 7.4, with HCl.…”