Persistent arterial hypotension is a hallmark of sepsis and is believed to be caused, at least in part, by excess nitric oxide (NO). NO can combine with superoxide to produce peroxynitrite, which activates matrix metalloproteinases (MMPs). Whether MMP inhibition in vivo protects against vascular hyporeactivity induced by endotoxemia is unknown. Male Sprague-Dawley rats were administered either bacterial lipopolysaccharide (LPS, 4 mg/kg ip) or vehicle (pyrogen-free water). Later (30 min), animals received the MMP inhibitor doxycycline (4 mg/kg ip) or vehicle (pyrogen-free water). After LPS injection (6 h), animals were killed, and aortas were excised. Aortic rings were mounted in organ baths, and contractile responses to phenylephrine or KCl were measured. Aortas and plasma were examined for MMP activity by gelatin zymography. Aortic MMP and inducible nitric oxide synthase (iNOS) were examined by immunoblot and/or immunohistochemistry. Doxycycline prevented the LPS-induced development of ex vivo vascular hyporeactivity to phenylephrine and KCl. iNOS protein was significantly upregulated in aortic homogenates from endotoxemic rats; doxycycline did not alter its level. MMP-9 activity was undetectable in aortic homogenates from LPS-treated rats but significantly upregulated in the plasma; this was attenuated by doxycycline. Plasma MMP-2 activities were unchanged by LPS. Specific MMP-2 activity was increased in aortas from LPS-treated rats. This study demonstrates the in vivo protective effect of the MMP inhibitor doxycycline against the development of vascular hyporeactivity in endotoxemic rats.