Receptor-mediated modulation of avian caecal muscle contraction by melatonin: role of tyrosine protein kinase

Melatonin receptors in the quail caecum were studied by 2[125I]iodomelatonin binding assay and the involvement of tyrosine protein kinase in the melatonin‐induced contraction was explored. The binding of 2[125I]iodomelatonin in the quail caecum membrane preparations was saturable, reversible and of high affinity with an equilibrium dissociation constant (Kd) of 24.6 ± 1.1 pm (n=7) and a maximum number of binding sites (Bmax) of 1.95 ± 0.09 fmol (mg/protein) (n=7). The relative order of potency of indoles in competing for 2[125I]iodomelatonin binding was: 2‐iodomelatonin > melatonin > 2‐phenylmelatonin > 6‐chloromelatonin > 6‐hydroxymelatonin > N‐acetylserotonin, indicating that ML1 receptors are involved. The binding was inhibited by Mel1b melatonin receptor antagonists, luzindole and 4‐phenyl‐2‐propionamidotetralin (4‐P‐PDOT) as well as by non‐hydrolyzable analogs of GTP like GTPγS and Gpp(NH)p but not by adenosine nucleotides. The latter suggests that the action of melatonin on the caecum is G‐protein linked. Cumulative addition of melatonin (1–300 nM) potentiated both the amplitude and frequency of spontaneous contractions in the quail caecum. The potentiation of rhythmic contractions was blocked by both luzindole and 4‐P‐PDOT. Antagonists of tyrosine kinase, genistein(2 μM) and erbstatin(4 μM) suppressed the modulation of spontaneous contractions by melatonin, but not inhibitors of protein kinase C (PKC) or protein kinase A (PKA). Melatonin‐induced increment in spontaneous contraction was blocked by nifedipine (0.4 nM). Thus, we suggest that melatonin potentiates spontaneous contraction in the quail caecum via interacting with G‐protein‐coupled Mel1b receptor which may activate L‐type Ca2+ channels by mobilizing tyrosine kinases.