5‐Methoxytryptamine Inhibits Cyclic AMP Accumulation in Cultured Retinal Neurons Through Activation of a Pertussis Toxin‐Sensitive Site Distinct from the 2‐[¹²⁵I]Iodomelatonin Binding Site

Abstract

Melatonin and 5‐methoxytryptamine inhibited forskolin‐stimulated cyclic AMP formation in cultured neural cells prepared from embryonic chick retina. Both methoxyindoles exhibited similar potency and efficacy, with EC50 values of 0.8 nM for melatonin and 7.2 nM for 5‐methoxytryptamine. Inhibition of cyclic AMP formation by 5‐methoxytryptamine or melatonin was prevented by pretreatment with pertussis toxin. Pretreatment of cultures with 5‐methoxytryptamine for 24 h reduced the subsequent inhibitory cyclic AMP response to 5‐methoxytryptamine but not that to 2‐iodomelatonin. Putative melatonin receptors on cultured retinal cells were labeled with 2‐[125I]iodomelatonin. Melatonin displaced specific 2‐[125I]iodomelatonin with a Ki value (0.8 nM) similar to the EC50 for inhibition of cyclic AMP formation. In contrast, 5‐methoxytryptamine only inhibited 2‐[125I]iodomelatonin binding at very high concentrations (Ki = 650 nM). Pretreating cultured cells for 24 h with 2‐iodomelatonin or melatonin, but not with 5‐methoxytryptamine, reduced subsequent 2‐[125I]iodomelatonin binding. Thus, 5‐methoxytryptamine appears to inhibit forskolin‐stimulated cyclic AMP formation at a site distinct from the 2‐iodomelatonin binding site.

Melatonin inhibitory effect on cAMP accumulation in the chick retina development

“…We showed that in the embryonic chick retinal cells in culture the nicotinic acetylcholine alpha‐bungarotoxin‐sensitive binding sites develop into functional receptors only in the presence of endogenous melatonin (Sampaio et al, 2005), an effect by diminishing the cAMP levels (Almeida‐Paula et al, 2005). The inhibition by melatonin of the intracellular cAMP levels in the embryonic chick retinal cells in culture already has been showed (Iuvone and Gan, 1994, 1995; Iuvone et al, 1995). Thus, a relationship between the melatonin inhibitory effect on total amount of cAMP and the modulation by melatonin of the neuronal differentiation is highly probable.…”

“…In this tissue, melatonin is related to modulation of dopamine release, activation of rod shedding, enhancing of horizontal cells sensitivity and modulation of the circadian electroretinogram rhythms (Dubocovich, 1985; Wiechmann et al, 1988; Nguyen‐Legros and Hicks, 2000; Peters and Cassone, 2005). The general mechanism surrounding the melatonin effects is the inhibition of the adenylate cyclase enzyme, directly by melatonin binding on G protein coupled receptors (Iuvone et al, 1995; Mackenzie et al, 2002; Dubocovich et al, 2003; Markowska et al, 2004), or indirectly by melatonin inhibiting the calmodulin enzyme, resulting in the inhibition of the group of the calmodulin‐dependent adenylate cyclase (Almeida‐Paula et al, 2005). Commonly, the melatonin receptors can also be related to other signal transduction cascades, depending on of the use of the native tissues or of the expression systems in the investigation, and of the melatonin concentration applied (Mackenzie et al, 2002; Dubocovich et al, 2003; Gerdin et al, 2003; Markowska et al, 2004; Schuster et al, 2005).…”

Melatonin, hormone of darkness and more – occurrence, control mechanisms, actions and bioactive metabolites

“…5-MT is a bioactive compound too [8,10,20] and may play a role of its own in the retina. In cultured retinal cells, it reportedly prevented the forskolin-induced rise in cyclic AMP, independently of the melatonin receptors [21]. Where not released from an organ, melatonin cannot not be degraded in the liver or appear as urinary 6-sulfatoxymelatonin.…”

Melatonin Beyond Its Classical Functions

“…[16], but the physiological relevance of respective pharmacological experiments is not always easy to judge. In cultured retinal cells, 5-MT was reported to prevent the forskolin-induced rise in cyclic AMP, in an action independent of melatonin receptors [94], a finding of still uncertain meaning.…”