Modulation of Conventional Outflow Facility by the Adenosine A1 Agonist N6-Cyclohexyladenosine

tigate the actions of the adenosine A1 agonist N 6 -cyclohexyladenosine (CHA) on conventional outflow facility. METHODS. Conventional outflow facility was evaluated in isolated bovine anterior segments, perfused at a constant pressure of 10 mm Hg. After overnight perfusion to establish a stable baseline, the concentration- and time-dependent changes in outflow facility induced by CHA were determined. To confirm the involvement of adenosine A1 receptors and matrix metalloproteinases (MMP) in any change in facility, the responses to CHA were evaluated in preparations treated with the adenosine A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT), or the nonselective MMP inhibitor GM-6001. RESULTS. The administration of CHA (10 M) to perfused anterior segments produced a 28% increase in outflow facility over basal levels. This response was relatively slow to develop with no significant change in outflow facility measured until after 60 minutes of CHA infusion. The peak response to CHA infusion occurred between 3 and 4 hours after CHA administration. Analysis of the CHA concentration‐response curves demonstrated that this increase in outflow facility was concentrationdependent, with an EC50 of 0.28 M. Pretreatment with the adenosine A1 receptor antagonist CPT (10 M) or the nonselective MMP inhibitor GM-6001 (10 M) blocked the response to CHA (1 M). When compared with control eyes, no significant change in baseline facility was measured in eyes perfused with CPT or GM-6001. CONCLUSIONS. These studies demonstrate that the adenosine agonist CHA significantly increases conventional outflow facility in the perfused bovine eye. Analysis of the CHA concentration‐response curve and inhibition of the CHA-induced increase in outflow facility by the adenosine A1 antagonist confirms that this response is mediated by the activation of adenosine A1 receptors. The inhibition of the CHA-induced increase in outflow facility by the MMP inhibitor GM-6001 provides evidence that the secretion and activation of MMPs within the conventional outflow pathway play a central role in the ocular hypotensive action of adenosine A1 agonists. (Invest Ophthalmol Vis Sci. 2005;46:3795‐3799) DOI:10.1167/ iovs.05-0421 I n primary open-angle glaucoma, elevated IOP results from increased resistance within the conventional outflow pathway. 1 Although several drugs have been developed to control this increase in IOP, most of these agents act on target tissues outside of the conventional outflow pathway. The identification of agents and their associated receptors that act on cells within the conventional pathway offer the potential to develop therapies that target the site responsible for the ocular hypertension observed in primary open-angle glaucoma. Adenosine is considered a reactive metabolite involved in cellular communication during periods of stress. In the eye, adenosine levels have been shown to increase during periods of retinal ischemia and elevated intraocular pressure. 2,3 Biochemical, pharmacological, and molecular studies have identified four adenosine receptor subtypes: A1 ,A 2a ,A 2b, and A3. 4 Recent in vitro studies have identified functional adenosine receptors on both trabecular meshwork and Schlemm’s canal cells. 5‐7 In vivo studies have shown that the administration of adenosine A1 receptor agonists lowers IOP in rabbits, mice, and monkeys. 8‐10 Although this reduction in IOP involves an early decrease in aqueous flow, most of this ocular hypotensive response results from an increase in total outflow facility. 10,11 In vitro studies have shown that adenosine A1 receptors are expressed by trabecular meshwork cells and the activation of these receptors are linked to the secretion of matrix metalloproteinases (MMPs). 6 However, studies have not been conducted to evaluate whether the activation of adenosine A1 receptors within the conventional outflow pathway contribute to the increase in total outflow facility. Moreover, the cellular events that mediate any functional changes in outflow have not been identified. To investigate whether adenosine A1 receptor activation modulates conventional outflow facility, we evaluated the effects of the adenosine A1 agonist N 6 -cyclohexyladenosine (CHA) on outflow facility in isolated perfused bovine anterior segments. Our results demonstrate that the administration of CHA increased outflow facility in this model. This increase in outflow facility was dependent on adenosine A1 receptor activation and blocked by pretreatment with the broad-spectrum MMP inhibitor GM-6001.