Effects of pinealectomy and melatonin on the retrograde degeneration of retinal ganglion cells in a novel model of intraorbital optic nerve transection in mice Introduction The retinotectal projection is a well-de®ned experimental system which permits the study of mechanisms of degen- eration of injured central nervous system (CNS) neurons in vivo. In adult mammalians, transection of the optic nerve (ON) close to the cell body results in delayed retrograde death of retinal ganglion cells (RGCs) (Garcia-Valenzuela et al., 1994; Isenmann et al., 1997). This model system has been used in recent years to study the eectiveness of neuroprotective strategies in rats (Isenmann et al., 1998; Kermer et al., 1998; KloÈcker et al., 1998). Until now, there was no comparable animal model available allowing to study RGC injury in mice. In recent years, mice have achieved increasing interest for the study of neurodegener- ation, as the genetic background of these animals is well-characterized and because an increasing number of transgenic and knockout animals have become available in which expression of speci®c regulatory proteins responsible for cell survival and death are modi®ed (Cellerino et al., 2000). Because of their small size, mice permit the inves- tigation of the eects of many pharmacological substances at reasonable expense, particularly when the amount of a substance is limited. Melatonin is a powerful free radical scavenger and anti- oxidant, which scavenges the highly toxic hydroxyl radical as well as other radicals that initiate lipid peroxidation (for review, see Reiter, 1998). As melatonin synthesis and secre- tion decrease markedly during aging (Reiter, 1992), mela- tonin de®ciency has been implicated in the pathophysiology of age-related neurodegenerative conditions, in which oxidative damage is involved. In fact, melatonin protects against delayed tissue damage after traumatic brain injury (Mesenge et al., 1998), focal cerebral ischemia (Manev et al., 1996; Kilic et al., 1999), excitotoxic brain lesions (Giusti et al., 1996, 1997; Manev et al., 1996; Uz et al., Kilic E, Hermann DM, Isenmann S, BaÈhr M. Eects of pinealectomy and melatonin on the retrograde degeneration of retinal ganglion cells in a novel model of intraorbital optic nerve transection in mice. J. Pineal Res. 2002; 32:106±111. ã Munksgaard, 2002 Abstract: The eects of pinealectomy and of intraperitoneally administered melatonin on the retrograde degeneration of retinal ganglion cells (RGCs) were examined in a novel model of optic nerve (ON) transection in C57BL/6J mice. RGCs were prelabeled with the ¯uorescent tracer 1,1¢-dioctadecyl- 3,3,3¢,3¢-tetramethyl indocarbocyanine perchlorate (Di-I), and the ON was cut inside the orbital cavity 7 days later. The degree of RGC injury was assessed by counting viable Di-I labeled RGCs in various locations of the retina. In unlesioned control eyes, a mean ganglion cell density of 1891  30/mm 2 (mean  S.E.M.) was determined. The cell density markedly declined at 14 days after axotomy (295  9 cells/mm 2 ; 15.6% of contralateral). Sham-pinealectomy did not in¯uence the density of RGCs at 14 days after ON transection (382  37 cells/mm 2 ). In pinealectomized animals, on the other hand, the RGC number was signi®cantly reduced as compared with untreated and sham-pinealectomized animals (91  33 RGCs/mm 2 ). The effect of pinealectomy was reversed after i.p. administration of melatonin (4 mg/kg bw bolus followed by continuous infusion of 8 mg/kg bw/day) (286  27 cells/mm 2 ). In nonpinealectomized animals, on the contrary, i.p. melatonin did not in¯uence the RGC density (344  20 cells/mm 2 ). The present results suggest that endogenous melatonin prevents the delayed degeneration of adult central nervous system (CNS) neurons in vivo, and that exogenous substitution of melatonin may be useful to protect injured neurons against cell death under conditions of melatonin de®ciency, e.g. in the aged brain, when melatonin synthesis and secretion have decreased. Ertugrul Kilic, Dirk M. Hermann, Stefan Isenmann and Mathias Ba Èhr Department of Neurology, University of Tu È bingen, Auf der Morgenstelle, Tu È bingen, Germany Key words: aging, antioxidant, melatonin, neurodegeneration, neuroprotection, pineal gland Address reprint requests to Dr Ertugrul Kilic, Department of Neurology, Georg August University, Robert-Koch-Str. 40, D-37075 Go È ttingen, Germany. E-mail: ertugrul.kilic@med.uni-goettingen.de Received April 19, 2001; accepted June 13, 2001. J. Pineal Res. 2002; 32:106±111 Copyright ã Munksgaard, 2002 Journal of Pineal Research ISSN 0742-3098 106