Characterization of functional VIP/PACAP receptors in the human erythroleukemic HEL cell line

Abstract

R. LEMA-KISOKA, N. HAYEZ, I. LANGER, P. ROBBERECHT, E. SARIBAN AND C. DELPORTE. Characterization of functional VIP/PACAP receptors in the human erythroleukemic HEL cell line. PEPTIDES. The presence of VIP/PACAP receptors was investigated on the human erythroleukemic cell line HEL. Specific binding of [ 125I]-PACAP or [ 125I]-VIP on HEL cells or membranes was very low and did not allow to perform competition curves. At 37°C PACAP transiently increased cAMP levels in the presence of the non-specific phosphodiesterase inhibitor IBMX, suggesting rapid desensitization. Kinetic studies revealed that optimal conditions to measure the EC 50 of PACAP(1–27) were 10 min at 20°C. Under those conditions, PACAP-related peptides increased cAMP levels with EC 50 in agreement with the pharmacological profile of the VPAC 1 receptor subtype: PACAP = VIP > [K 15, R 16, L 27]VIP(1–7)/GRF(8–27) = [R 16]ChSn (two VPAC 1 agonists) ≫ helodermin = secretin. RO 25–1553, a selective activator of VPAC 2 receptor was inactive at 1 μM. Dose-response curves of VPAC 1 agonist molecules (PACAP, VIP, [K 15, R 16, L 27]VIP(1–7)/GRF(8–27), [R 16]ChSn) were shifted to the right by the VPAC 1 receptor antagonist [AcHis 1, D-Phe 2, Lys 15, Leu 17]VIP(3–7)/GRF(8–27), with a K i of 3 ± 1 nM ( n = 3). The presence of VPAC 1 receptor mRNA was confirmed by RT-PCR. Preincubation with PACAP or PMA showed that VPAC 1 receptors underwent homologous and heterologous desensitization. This study provides the first evidence for the expression of functional VPAC 1 receptors undergoing rapid desensitization in HEL cells.