Cloning and expression of a complementary DNA encoding a molluscan octopamine receptor that couples to chloride channels in HEK293 cells.
A cDNA encoding a G-protein-coupled receptor was cloned from the central nervous system of the pond snail Lymnaea stagnalis. The predicted amino acid sequence of this cDNA most closely resembles the Drosophila tyramine/octopamine receptor, the Locusta tyramine receptor, and an octopamine receptor (Lym oa1) that we recently cloned from Lymnaea. After stable expression of the cDNA in HEK293 cells, we found that [3H]rauwolscine binds with high affinity to the receptor (KD = 6.2.10(-9) M). Octopamine appears to be the most potent naturally occurring agonist to displace the [3H]rauwolscine binding (Ki = 3.0.10(-7) M). Therefore, the receptor is considered to be an octopamine receptor and is consequently designated Lym oa2. The novel receptor shares little pharmacological resemblance with Lym oa1, indicating that the two receptors represent different octopamine receptor subfamilies. Octopaminergic stimulation of Lym oa2 does not induce changes in intracellular concentrations of cAMP or inositol phosphates. However, electrophysiological experiments indicate that octopamine is able to activate a voltage-independent Cl- current in HEK293 cells stably expressing Lym oa2. Although opening of this chloride channel most probably does not require the activation of either protein kinase A or C, it can be blocked by inhibition of protein phosphorylation.
Gerhardt
C C
CC
Department of Biochemistry and Molecular Biology, Research Institute Neurosciences, Vrije Universiteit, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
Lodder
H C
HC
Vincent
M
M
Bakker
R A
RA
Planta
R J
RJ
Vreugdenhil
E
E
Kits
K S
KS
van Heerikhuizen
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D018118:Chloride Channels / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D019204:GTP-Binding Proteins / Q000502:physiology; D006801:Humans; D015640:Ion Channel Gating; D008195:Lymnaea / Q000737:chemistry / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000235:genetics; D008969:Molecular Sequence Data; D009655:Octopamine / Q000502:physiology*; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D017982:Receptors, Biogenic Amine / Q000378:metabolism*; D015398:Signal Transduction; D015016:Yohimbine / Q000378:metabolism
D018118:Chloride Channels; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D017982:Receptors, Biogenic Amine; C027604:norsynephrine receptor; D009655:Octopamine; D015016:Yohimbine; D019204:GTP-Binding Proteins