Many G protein coupled receptors (GPCRs) can be activated by neurotransmitters, acting via heteromeric G proteins to control downstream intracellular processes. A GPCR can be simplified as either excitatory (Galphas or Galphaq) or inhibitory (Galphai/o). While ligands for many C. elegans neurotransmitter GPCRs have been identified, many gaps in our knowledge remain. To address this, we expressed C. elegans GPCRs in Xenopus oocytes along with G protein-coupled inwardly rectifying potassium channels (GIRKs) to evaluate ligand specificity and G-protein coupling using two-electrode voltage clamp recordings. Using this approach, we have successfully observed activation of 10 monoamine-activated GPCRs, all with previously identified ligands. Despite this previous knowledge, some results on their G-protein coupling, and the potency of secondary ligands was often conflicting or missing. For example, SER-6 and DOP-4 have previously been shown to couple to Galphas but our experiments indicate Galphaq coupling. Activation by two or more ligands was observed for all the GPCRs tested, but the published data does not reflect this phenomenon. Several additional orphan receptors were evaluated, DOP-5, DOP-6, SER-5 and PCDR-1, and no activation was detected. The reason is unknown, but it could be because they require a GPCR partner to form a functional receptor, because they couple to a non-cannonical G-protein, or because we have not tested the correct ligands. In addition, we have characterized the putative adenosine receptor ADOR-1 and found it to be indeed activated by adenosine. Surprisingly, we obtained evidence suggesting coupling to the excitatory Galphaq protein as well as to Galphai/o or Galphas. Further experiments are ongoing to confirm this dual coupling in vitro and in vivo. Using endogenous fluorescent reporters, we found
ador-1 to be broadly expressed in the nervous system as well as in muscle, including MC/I4/pharyngeal muscle and vulval muscle/HSN. However, despite this expression pattern we did not observe any significant differences in pumping rate or egg laying under normal conditions. In other species, intracellular adenosine levels have been shown to increase after injury or stress, and we observed broad expression of
ador-1 in multiple 'avoidance' neurons (AWB/ASH/ASK). We hypothesise that adenosine may act via
ador-1 to regulate stress related behaviours or lifespan and we plan to investigate this going forward.