Orexin Receptor Type-1 Couples Exclusively to Pertussis Toxin-Insensitive G-Proteins, While Orexin Receptor Type-2 Couples to Both Pertussis Toxin-Sensitive and -Insensitive G-Proteins

Abstract

Signal transduction pathways of orexin receptors were examined using a nerve-like cell line transfected with orexin receptor type-1 (OX1R) and orexin receptor type-2 (OX2R). Forskolin-stimulated cyclic adenosine 3,5-monophosphate (cAMP) accumulation in OX2R-expressing cells was inhibited by orexin in a dose-dependent manner, and the effect was abolished by pretreatment with pertussis toxin (PTX). The inhibitory effect of orexin on forskolin-stimulated cAMP accumulation was not observed in OX1R-expressing cells. Administration of orexin to these cells resulted in a transient increase of intracellular calcium concentration ([Ca(2+)](i)). Orexin-stimulated increases in [Ca(2+)](i) in OX1R- or OX2R-expressing cells were not affected by the PTX pretreatment. These observations suggest that OX1R couples exclusively to PTX-insensitive G-proteins, while OX2R couples to both PTX-sensitive and -insensitive G-proteins. To examine the relative contributions of these G-proteins in OX2R-mediated activation of neurons, we used histaminergic tuberomammillary nucleus neurons, in which OX2R is abundantly expressed. We found that a phospholipase C (PLC)-inhibitor, U73122, inhibits orexin-mediated neuronal activation, but PTX showed no effect on it. This suggests that although OX2R couples to multiple G-proteins, activation of neurons by orexins through OX2R is mediated via a PTX-insensitive, PLC dependent pathway.

Orexin Directly Excites Orexin Neurons through Orexin 2 Receptor

“…This suggests the activation of nonselective cation channels downstream of OX2R. The activation of phospholipase C by the ␤␥ subunit of the G-protein might be involved because OX2R is believed to couple with the Gq and Gi types of G-proteins (Zhu et al, 2003). We previously showed that orexin neurons are activated by CCK and AVP via the G-protein-coupled receptors (Gq), CCK A receptor, and the V1a receptor (Tsujino et al, 2005;Tsunematsu et al, 2008).…”

The Orexin OX1Receptor Regulates Ca2+Entry via Diacylglycerol-Activated Channels in Differentiated Neuroblastoma Cells

“…This is frequently observed in heterologous expression systems with both OX 1 R and OX 2 R (Smart et al, 1999;Holmqvist et al, 2002;Ammoun et al, 2003) and is likely to be a consequence of activation of phospholipase C (PLC) and the generation of IP 3 . Some studies on native receptors in neurons have demonstrated sensitivity to inhibitors of phosphatidylinositol-specific PLC (Zhu et al, 2003;Muroya et al, 2004), suggesting that some store release may take place. Conversely, a Ca 2ϩ release from intracellular stores has not been demonstrated in native neurons although explicitly tested for Kohlmeier et al, 2004).…”

Orexin in Rostral Hotspot of Nucleus Accumbens Enhances Sucrose ‘Liking’ and Intake but Scopolamine in Caudal Shell Shifts ‘Liking’ Toward ‘Disgust’ and ‘Fear’

“…Orexin is typically thought to have excitatory depolarization effects on neurons (Korotkova et al, 2003;Marcus et al, 2001;Sakurai et al, 1998;Trivedi et al, 1998;van den Pol et al, 2002;Zhu et al, 2003). However, neurons in NAc may exclusively contain orexin-2 receptors (OX r 2) (Ch'ng and Lawrence, 2015; Trivedi et al, 1998), which can be coupled with either Gi or Gq subunits, and may inhibit neurons via the augmentation of inhibitory GABA signals (Martin et al, 2002;Zhu et al, 2003). If so, neuronal inhibition by orexin could be more similar to GABAergic hyperpolarizations or Gi-coupled opioid or endocannabinoid signaling.…”