To the Editor:
The paper of Espino et al. (1) addressed a very interesting issue about the anti-apoptotic activity of the pineal hormone melatonin in human semen. Employing 1 mM doses of melatonin in vitro and the MT1/MT2 receptor antagonists luzindole and 4-phenyl-2-propionamidotetralin (4P-PDOT), the authors claim to have demonstrated that “…melatonin prevention of H2O2-induced DNA fragmentation is dependent on …MT1 receptor ….”. We find that above statement may be arguable and there is need of further exploration and clarification of the mechanisms involved.
Protection against oxidative damage and DNA fragmentation by melatonin has been a matter of investigation in various experimental models. Unfortunately, as in the case with the design used by Espino et al. (1), the results from many in vitro studies were difficult to interpret because experiments were performed with extremely high concentrations of melatonin, which would never be reached under physiological conditions of circadian-timed release of the pineal hormone or after most therapeutic applications. High nocturnal melatonin levels in humans (100–400 pM) are considered to be saturating for the high-affinity, low-capacity MT1/MT2 receptors, given the potency of melatonin established in ligand-binding experiments (2, 3). Single reports of melatonin concentration in human semen indicate low levels, between 2.6 and 21 pM (4), which seem reasonable for the activation of putative membrane receptors on human spermatozoa. Given that the known G protein-coupled membrane melatonin receptors MT1 and MT2 are activated by picomolar concentrations of melatonin, it is not plausible that specific membrane receptor-dependent signaling events can be triggered by 3- to 6-fold higher ligand concentrations. Considering the above points, the results from the dose-response experiments on the H2O2-induced caspase activation using high pretreatment melatonin doses should be interpreted with caution. Under conditions of concentration overload with melatonin, one should account for desensitization of the affinity-binding sites, which may have influenced the expression of the response. With this background, the use of luzindole and 4P-PDOT cannot warrant their clear antagonistic activity, given the pharmacologic profile of the drugs on recombinant and native melatonin receptors (2).
Early experience with melatonin from this laboratory (3) demonstrated that the in vitro use of the indole in doses higher than micromolar might not be justified in attempts to demonstrate membrane receptor-mediated action. According to other reports (5), melatonin’s effect on motility characteristics of ejaculated human spermatozoa can be observed in vitro by low nanomolar doses, and this might correspond to the expression of the membrane melatonin receptor. One alternative explanation for the observed effects in the study of Espino et al. (1) may be that the in vitro conditions used allowed for melatonin to influence membrane lipid peroxidation reactions and/or mitochondrial membrane permeability, with subsequent shifts in oxidative species-induced caspase activation.
Rossitza Konakchieva, Ph.D., D.Sci.
Plamen Todorov, Ph.D.
Institute of Biology and Immunology of Reproduction
Bulgarian Academy of Sciences
1. Espino J, Ortiz A, Bejarano I, Lozano GM, Monllor F, Garc JF, Rodrıguez AB, Pariente JA. Melatonin protects human spermatozoa from apoptosis via melatonin receptor– and extracellular signal–regulated kinase-mediated pathways. Fertil Steril 2011;95:2290–6.
2. Dubocovich M, Delagrange Ph, Krause D, Sugden D, Cardinali D, Olcese J. International Union of Basic and Clinical Pharmacology. LXXV. Nomenclature, Classification, and Pharmacology of G Protein-Coupled Melatonin Receptors. Pharmacol Rev 2010;62:343–80.
3. Konakchieva R, Kyurkchiev S, Kehayov I, Taushanova P, Kanchev L. Selective effect of methoxyindoles on the lymphocyte proliferation and melatonin binding to activated human lymphoid cells. J Neuroimmunol 1995;63:125-32.
4. Luboshitzky R, Shen-Orr Z, Herer P. Seminal plasma melatonin and gonadal steroids concentrations in normal men. Arch Androl 2002;48:225-32.
5. Logoglu G, Kendirci A, Ilbay G. Melatonin exerts stimulatory effect on human sperm motility, in vitro. J Islam Acad Sci 1996;9:13-18.
Published online in Fertility and Sterility