Quantitative and regional measurement of retinal blood flow in rats using N-isopropyl-p-[ 14 C]-iodoamphetamine ([ 14 C]-IMP) Myle ` ne Pouliot a, b , Micheline C. Desche ˆnes a, c, d , Simon He ´ tu a, b , Sylvain Chemtob e , Mark R. Lesk c, d , Re ´ jean Couture b , Elvire Vaucher a, * a School of Optometry, University of Montreal, C.P. 6128, succursale Centre-Ville, Montre ´al, QC H3C 3J7, Canada 1 b Department of Physiology, University of Montreal, C.P. 6128, succursale Centre-Ville, Montre ´al, QC H3C 3J7, Canada 2 c Department of Ophthalmology, University of Montreal, C.P. 6128, succursale Centre-Ville, Montre ´al, QC H3C 3J7, Canada 3 d Research Centre Guy-Bernier, University of Montreal, Maisonneuve-Rosemont Hospital, 5415 boul. L’Assomption, Montre´al, QC H1T 2M4, Canada 4 e Departments of Pediatrics, Ophthalmology and Pharmacology, Research Centre of Sainte-Justine Hospital, 3175 Co ˆte Sainte-Catherine, Montre ´al, Que ´bec H3T 1C5, Canada 5 article info Article history: Received 9 June 2009 Accepted in revised form 12 August 2009 Available online 19 August 2009 Keywords: blood flow retina brain autoradiography hypercapnia iodoamphetamine microcirculation imaging abstract Quantitative and regional measurement of retinal blood flow in rodents is of prime interest for the investigation of regulatory mechanisms of ocular circulation in physiological and pathological conditions. In this study, a quantitative autoradiographic method using N-isopropyl-p- 14 C-iodoamphetamine ([ 14 C]-IMP), a diffusible radioactive tracer, was evaluated for its ability to detect changes in retinal blood perfusion during hypercapnia. Findings were compared to cerebral blood flow values measured simul- taneously. Hypercapnia was induced in awaken Wistar rats by inhalation of 5% or 8% CO 2 in medical air for 5 min. [ 14 C]-IMP (100 mCi/kg) was injected in the femoral vein over a 30 s period and the rats were sacrificed 2 min later. Blood flow was calculated from whole-mount retinae and 20 mm thick brain sections in discrete regions of interest by quantitative autoradiography or from digested samples of retina and brain by liquid scintillation counting. Retinal blood flow values measured with quantitative and regional autoradiography were higher in the central (108  20 ml/100 g/min) than in peripheral (84  15 ml/100 g/ min) retina. These values were within the same range as cortical blood flow values (97  4 ml/100 g/min). The retinal blood flow values obtained on whole-mount retinae were validated by the sampling method. Hypercapnia significantly increased overall blood flow in the retina (24–53%) with a maximal augmen- tation in the peripheral region and in the brain (22–142%). The changes were stronger in the brain compared to retina (p ¼ 0.016). These results demonstrate that retinal blood flow can be quantified using [ 14 C]-IMP and compared with cerebral blood flow. This technique is a powerful tool to study how retinal blood flow is regulated in different regions of the rat retina. Ó 2009 Elsevier Ltd. All rights reserved. 1. Introduction Deficit in blood supply in the retina contributes to the develop- ment of retinal diseases, such as diabetic retinopathy, age-related macular degeneration and glaucoma (Grunwald et al., 1984; Langham et al., 1991; Atmaca et al., 1996). Rodent models are commonly used to study the development of these retinal damages and to test potential therapies. However, blood flow dysfunctions in rodents and their cellular and molecular mechanisms remain elusive. This is principally due to a lack of a technical approach which would adequately assess all the specific features of the rodent retinal microcirculation. Due to its small size, laminar structure, location and apposition to the choroid, the rodent retina is particularly difficult to study. Optically based imaging techniques, including laser Doppler flowmetry (Tsuji- kawa et al., 2000; Yu et al., 2005; Chauhan et al., 2006; Mori et al., 2007), on-line video angiography (Clermont et al., 1994; Kunisaki et al., 1998) or optical coherence tomography (Fujimoto et al., 1995) have a great temporal resolution but a poor spatial and laminar resolution and require a high transparency of the light paths (Glazer, 1988; Duong et al., 2008). Alternatively, quantitative techniques such as the use of systemic radioactive or non-radioactive microspheres (Chemtob et al., 1991; Alm et al., 1997; Wang et al., 2007) and magnetic Abbreviations: CBF, cerebral blood flow; DLG, dorsolateral geniculate nucleus; IMP, iodoamphetamine; RBF, retinal blood flow. * Corresponding author. Tel.: þ1 514 343 7537; fax: þ1 514 343 2382. E-mail address: elvire.vaucher@umontreal.ca (E. Vaucher). 1 Tel.: þ1 514 343 7537; fax: þ1 514 343 2382. 2 Tel.: þ1 514 343 7060; fax: þ1 514 343 2111. 3 Tel.: þ1 514 343 7094; fax: þ1 514 343 5790. 4 Tel.: þ1 514 252 3400x4959; fax: þ1 514 343 3821. 5 Tel.: þ1 514 345 4692; fax: þ1 514 345 4801. Contents lists available at ScienceDirect Experimental Eye Research journal homepage: www.elsevier.com/locate/yexer 0014-4835/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.exer.2009.08.005 Experimental Eye Research 89 (2009) 960–966