Selective expression of vasoactive intestinal peptide (VIP)2/pituitary adenylate cyclase-activating polypeptide (PACAP)3 receptors in rabbit and guinea pig gastric and tenia coli smooth muscle cells.
In both functional and radioligand binding studies of gastric smooth muscle from rabbit and guinea pig, vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) show equal potency indicating that the receptor type is either a VIP1/PACAP2 or a VIP2/PACAP3 receptor. We have characterized the VIP/PACAP receptor expressed in freshly dispersed and cultured gastric and tenia coli smooth muscle cells of rabbit and guinea pig by reverse transcriptase-polymerase chain reaction (RT-PCR), Northern analysis, and cloning of the first extracellular domain. Specific primers based on cDNA sequences for rat VIP1/PACAP2, VIP2/PACAP3 and PACAP1 receptors were designed spanning the first extracellular domain. A 275 base pair product corresponding to VIP2/PACAP3 receptor was amplified by RT-PCR in muscle cells from both species. No RT-PCR product was obtained with primers for VIP1/PACAP2 and PACAP1 receptors. The deduced amino acid sequences showed 90% similarity in rabbit and 77% in guinea pig to the sequence in rat. The location of the aspartate, tryptophan and glycine residues and all six N-terminal cysteines required for VIP binding were conserved. The sequence in guinea pig tenia coli differed from that in guinea pig stomach by two amino acid residues, Phe40 and Phe41. Northern analysis revealed a single 3.9 kilobase (kb) mRNA corresponding to VIP2/PACAP3 receptors in rabbit and a 2.1 kb mRNA in guinea pig gastric and tenia coli muscle cells. We conclude that only VIP2/PACAP3 receptors are expressed in smooth muscle cells of rabbit and guinea pig. The two amino acid difference in the sequence obtained from guinea pig tenia coli may reflect the distinct binding and functional properties of this tissue.
Teng
B
B
Department of Physiology, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0711, USA.
Murthy
K S
KS
Kuemmerle
J F
JF
Grider
J R
JR
Makhlouf
G M
GM
D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003106:Colon / Q000378:metabolism*; D005753:Gastric Mucosa / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics*; D006168:Guinea Pigs; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D011979:Receptors, Pituitary Hormone / Q000235:genetics*; D018005:Receptors, Vasoactive Intestinal Peptide / Q000235:genetics*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D012333:RNA, Messenger; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D011979:Receptors, Pituitary Hormone; D018005:Receptors, Vasoactive Intestinal Peptide