Evidence that Dopamine is the Physiological Agonist of Spinal Ghrelin Receptors and Acts Through Heteromeric Dopamine D2/ Ghrelin Receptors

Ghrelin receptors (GhrR) occur in autonomic control centers in the spinal cord. However, the only natural activator of GhrR, ghrelin, is absent from the spinal cord. Therefore, we sought another explanation for the conservation of spinal GhrR across species and the consistency of stimulatory effects of GhrR agonists on autonomic centers in the spinal cord.We found that agonists of dopamine 2 receptors (DRD2) applied to spinal defecation centers in the rat mimic actions of ghrelin receptor agonists to stimulate colorectal propulsion and defecation. Here we present data indicating that the effects through spinal GhrR receptors are mediated by dopamine acting at DRD2 that is coupled to GhrR. In rats, we activated the defecation pathways by a natural stimulus, water avoidance stress, which caused increased fecal output in the rats. The responses were inhibited both by the DRD2 antagonist, sulpiride, and by the GhrR antagonist, YIL781, applied systemically. The stimulation of colorectal propulsion by the DRD2 agonist, quinpirole, applied directly to the defecation centers, was also reduced by GhrR antagonism with YIL781 (systemic). Colorectal effects of DRD2 and GhrR agonists were prevented by cutting the pelvic nerves, indicating that their sites of action are in the CNS. We used RNAScope to localize sites of DRD2 and GhrR expression in the spinal cord. DRD2 and GhrR mRNA were colocalized in autonomic preganglionic neurons, identified by their expression of choline acetyltransferase, at the levels of the defecation centers. Dual localization of the receptors occurred in the majority of ChAT neurons in both the intermediolateral (IML) and intermediomedial (IMM) autonomic cell groups. ChAT, DRD2 and GhrR mRNAs were expressed at similar levels in the neurons. In HEK cells transfected with DRD2, dopamine receptor agonists inhibited adenylyl cyclase but had no effect on cytoplasmic Ca2+. GhrR agonists had no effect in DRD2 cells. In cells transfected with DRD2 plus GhrR, agonists of either receptor evoked increases in cytoplasmic Ca2+. The effects were antagonized by the GhrR antagonist, YIL781, in both cases. We conclude that the involvement of spinal GhrR in colorectal control in the absence of ghrelin, that has been demonstrated both in human and in animal models, is explained by dopamine being the transmitter that acts at DRD2/ GhrR heteromeric receptorsSupport or Funding InformationFlorey Institute of Neuroscience and mental health