Transient ischemia of the retina results in massive degeneration of the retinotectal projection: long-term neuroprotection with brimonidine Marcelino Avile ´s-Trigueros, a Sergio Mayor-Torroglosa, b Antonio Garcı ´a-Avile ´s, b Marı ´a P. Lafuente, b Marı ´a E. Rodrı ´guez, b Jaime Miralles de Imperial, b Marı ´a P. Villegas-Pe ´rez, b and Manuel Vidal-Sanz b, * a Departamento de Histologı ´a, Instituto de Bioingenierı ´a, Facultad de Medicina, Universidad Miguel Herna ´ndez, Alicante, Spain b Departamento de Oftalmologı ´a, Facultad de Medicina, Universidad de Murcia, Murcia, Spain Received 26 March 2003; revised 21 May 2003; accepted 2 June 2003 Abstract In adult rats, we have induced retinal ischemia and investigated anterogradely labeled surviving retinal ganglion cell (RGC) afferents to the contralateral superior colliculus (SC). The animals received topically in their left eyes two 5-Al drops of saline or saline-containing 0.5% brimonidine (BMD), 1 h before 90 min of retinal ischemia induced by ligature of the left ophthalmic vessels. Two months after ischemia, the anterogradely transported neuronal tracer cholera toxin B subunit (CTB) was injected in the ischemic eyes and animals were processed 4 days later. As controls and for comparison, the retinotectal innervation of unlesioned age-matched control rats was also examined with CTB. In control and experimental animals, serial coronal sections of the mesencephalon and brainstem were immunoreacted for CTB and the area and thickness of the two most superficial layers of the SC containing densely CTB-labeled profiles were estimated with an image analysis system. Ninety minutes of ischemia resulted 2 months later in reduced density of CTB-labeled profiles in the contralateral SC of the vehicle-treated rats, representing less than one half the area occupied by CTB-labeled profiles in control rats. This resulted in shrinkage of these layers and in the presence of areas virtually devoid of CTB immunoreactivity, suggesting orthograde degeneration of retinal terminals and/or decrease of anterograde axonal transport. Topical pretreatment with BMD resulted 2 months later in CTB immunoreactivity that occupied the superficial layers of the contralateral SC in an area of approximately 86% of that observed in the unlesioned control group of animals, indicating that BMD protects against ischemia-induced degeneration of the retinotectal projection, and preserves anterograde axonal transport. D 2003 Elsevier Inc. All rights reserved. Keywords: Retinal ischemia; Neuroprotection; Orthograde tracers; Brimonidine; Retinotectal system; Alpha-2 selective agonists; Superior colliculus; Anterograde degeneration; Anterograde axonal transport; Retinal ganglion cell axons; Cholera toxin B subunit Introduction In mammals, visual information processed in the retina is conveyed to retinorecipient nuclei in the brain by retinal ganglion cell (RGC) axons. In the rat, the main retinoreci- pient target region in the brain is the superior colliculus (SC) or optic tectum, where virtually all RGCs project (Lund, 1965). This structure is involved in the generation of integrated responses to visual, auditory and somatosensory stimuli (reviewed in Sefton and Dreher, 1985). The SC is a bilateral structure of the brainstem that contains several layers; the superficial layers, also known as upper or visual layers, receive their inputs from retinal and primary visual cortex afferents and comprise the stratum zonale (SZ), stratum griseum superficiale (SGS) and stratum opticum (SO) (Huber and Crosby, 1943). In the SC, RGC axons are deployed in a very precise topographic ordered manner (Linden and Perry, 1983; Sauve ´ et al., 2001, 2002). The retinotectal pathway is a suitable region of the central nervous system to investigate the effects of transient ischemia of the retina as well as to develop strategies toward its prevention (Vidal-Sanz et al., 2003). Using retrogradely transported neuronal tracers to identify the RGC population, we have recently shown that transient ischemia of the retina induces RGC loss, whose severity and duration is related to the initial ischemic interval (Lafuente et al., 2001, 2002a). Furthermore, a proportion of the RGC population that 0014-4886/$ - see front matter D 2003 Elsevier Inc. All rights reserved. doi:10.1016/S0014-4886(03)00298-X * Corresponding author. Departamento de Oftalmologı ´a, Facultad de Medicina, Universidad de Murcia, Campus de Espinardo, E-30.100 Espi- nardo, Murcia, Spain. Fax: +34-968-363962. E-mail address: ofmmv01@um.es (M. Vidal-Sanz). www.elsevier.com/locate/yexnr Experimental Neurology 184 (2003) 767 – 777