br Acknowledgments The authors thank Elisabete Lepera and Ro
Acknowledgments The authors thank Elisabete Lepera and Rosana Silva for technical support. The study was supported by FAPESP, CNPq and PADC/FCF-UNESP. T.T. Miguel was the recipient of a FAPESP (Proc. 05/05171-1) and R.L. Nunes-de-Souza of a CNPq (Proc. 303580/2009-7) fellowship.
Introduction Corticotropin-releasing factor (CRF) is a 41 amino potassium hydrochloride peptide that plays an important role in the regulation of the hypothalamic–pituitary–adrenal axis and in endocrine, behavioral, and autonomic responses to stress (Koob and Heinrichs, 1999). CRF belongs to a family of structurally related peptides that includes urocortin 1 (Vaughan et al., 1995), urocortin 2 (Reyes et al., 2001), and urocortin 3 (Lewis et al., 2001), that have been identified in humans, rodents, and other mammalian species. In addition to effects on the pituitary and the central nervous system (CNS), CRF and related peptides have been shown to modulate a range of peripheral activities in mammals including cardiovascular and gastrointestinal functions and inflammatory processes. CRF-related peptides are known to be the endogenous mammalian ligands for two G-protein coupled receptor subtypes, CRF1 and CRF2, that are positively coupled to adenylate cyclase (Perrin and Vale, 1999). The CRF2 receptor subtype has two functional splice variants in rodents, CRF2(a) and CRF2(b), which are approximately 70% identical in amino acid sequence Lovenberg et al., 1995, Perrin and Vale, 1999. The two CRF receptors differ in their pharmacological profiles. Urocortin 1, urocortin 2, and urocortin 3 bind to the CRF2 subtype with much greater affinity than CRF itself Lewis et al., 2001, Perrin and Vale, 1999, Reyes et al., 2001. In contrast, urocortin 2 and urocortin 3 show no appreciable binding to the CRF1 subtype Lewis et al., 2001, Reyes et al., 2001, whereas CRF and urocortin 1 bind to the CRF1 receptor with similar potency Perrin and Vale, 1999, Vaughan et al., 1995. Urocortin 2 is a selective CRF2 receptor peptide agonist with a Ki value of 0.66 nM for inhibition of radiolabeled sauvagine binding to recombinant mouse CRF2(b) receptors and >100 nM for the recombinant human CRF1 receptor (Reyes et al., 2001). Antisauvagine-30 is a selective CRF2 receptor peptide antagonist, with reported Ki values for inhibition of radiolabeled sauvagine binding to recombinant mouse CRF2(b) and rat CRF1 receptors of 1.4 and 154 nM, respectively, at room temperature (Rühmann et al., 1998) and a Kd value for radiolabeled antisauvagine-30 binding to recombinant CRF2(a) receptors of 0.125 nM with no specific binding detected to the recombinant human CRF1 receptor (Higelin et al., 2001). The discovery of urocortin 2 and the advent of the selective CRF2 receptor antagonist antisauvagine-30 have provided the pharmacological tools to explore the physiological relevance of the CRF2 receptor subtype in vivo (Pelleymounter et al., 2002). There is evidence available to suggest that modulation of CRF2 receptors can have effects on cardiovascular function in rodents (Coste et al., 2002). The CRF2 receptor subtype is localized in cardiac tissue and in the peripheral vasculature of normal animals Baigent and Lowry, 2000, Kishimoto et al., 1995, Lovenberg et al., 1995. Systemic administration of urocortin 1 has been shown to induce a marked and persistent reduction in blood pressure, putatively through activation of peripheral CRF2 receptors Lawrence et al., 2002, Spina et al., 1996, Vaughan et al., 1995. In addition, CRF2-deficient mice show elevated mean arterial blood pressure and diastolic pressure compared with wild-type littermates Bale et al., 2000, Coste et al., 2000. Moreover, the levels of mRNA for the CRF2 receptor in the cardiac tissue of spontaneously hypertensive rats are significantly higher than those in normotensive animals (Makino et al., 1998). Collectively, these data suggest that the CRF2 receptor subtype may play a role in modulation of the cardiovascular system. In order to more precisely address this question, we have investigated the effects of systemic administration of the selective CRF2 receptor peptide agonist urocortin 2 and the selective CRF2 receptor peptide antagonist antisauvagine-30 on mean arterial blood pressure and heart rate in conscious, freely moving rats.