A putative tachykinin receptor in the cockroach brain:: molecular cloning and analysis of expression by means of antisera to portions of the receptor protein

Tachykinins constitute a neuropeptide family that mediate their actions via a subfamily of structurally related G-protein-coupled receptors. Two receptors, Drosophila neurokinin receptor (NKD) and Drosophila tachykinin receptor (DTKR), with sequence similarities to mammalian tachykinin receptors have previously been cloned in Drosophila. In this study we have isolated a cockroach (Leucophaea maderae) cDNA clone by screening a brain cDNA library with a degenerate oligonucleotide probe based on a conserved sequence within the seventh transmembrane region of the Drosophila tachykinin receptors. This clone, Leucophaea tachykinin receptor (LTKR), encodes a portion of a putative receptor which could be aligned with the C-terminal half of members of the tachykinin receptor subfamily. In the fifth, sixth and seventh transmembrane regions the deduced amino acid sequence of LTKR exhibits 79% sequence identity to the DTKR receptor and 54%, to that of NKD. This suggests that LTKR is orthologous to the DTKR receptor. To study the distribution of the predicted LTKR protein by immunocytochemistry, antisera were raised against synthetic peptides corresponding to a region of the, third intracellular loop of LTKR. In the cockroach brain immunoreactive neuronal processes were seen in several synaptic neuropils of the protocerebrum and tritocerebrum, as well as in the frontal ganglion. Some immunoreactive neuronal cell bodies were detected in the protocerebrum. Double labeling immunocytochemistry revealed that there is a substantial superposition between distribution of LTKR and processes containing tachykinin-related peptide (TRP). Some brain areas, however, only display TRP immunoreactive processes and no LTKR, suggesting the presence, of at least one more TRP receptor type. (C) 2001 Elsevier Science BY All rights reserved.