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 recep tors, Drosophila neurokinin receptor (NKD) and Drosophila tachykinin recept or (DTKR), with sequence similarities to mammalian tachykinin receptors hav e previously been cloned in Drosophila. In this study we have isolated a co ckroach (Leucophaea maderae) cDNA clone by screening a brain cDNA library w ith a degenerate oligonucleotide probe based on a conserved sequence within the seventh transmembrane region of the Drosophila tachykinin receptors. T his clone, Leucophaea tachykinin receptor (LTKR), encodes a portion of a pu tative receptor which could be aligned with the C-terminal half of members of the tachykinin receptor subfamily. In the fifth, sixth and seventh trans membrane regions the deduced amino acid sequence of LTKR exhibits 79% seque nce identity to the DTKR receptor and 54%, to that of NKD. This suggests th at LTKR is orthologous to the DTKR receptor. To study the distribution of t he predicted LTKR protein by immunocytochemistry, antisera were raised agai nst synthetic peptides corresponding to a region of the, third intracellula r loop of LTKR. In the cockroach brain immunoreactive neuronal processes we re seen in several synaptic neuropils of the protocerebrum and tritocerebru m, as well as in the frontal ganglion. Some immunoreactive neuronal cell bo dies were detected in the protocerebrum. Double labeling immunocytochemistr y revealed that there is a substantial superposition between distribution o f LTKR and processes containing tachykinin-related peptide (TRP). Some brai n areas, however, only display TRP immunoreactive processes and no LTKR, su ggesting the presence, of at least one more TRP receptor type. (C) 2001 Els evier Science BY All rights reserved.