1. In recent studies, the vascular adventitia has been established as an important source of inducible nitric oxide synthase (iNOS) and subsequent nitric oxide (NO) production, even more powerful than the media in response to certain inflammatory factors, such as lipopolysaccharide (LPS). The adventitia has an independent L-arginine (L-Arg)/NOS/NO pathway and is involved in the regulation of vascular function. In the present study, we explored the changes in and the pathophysiological significance of the L-Arg/NOS/NO pathway in the adventitia of rats with sepsis. 2. Sepsis was induced by caecal ligation and puncture in order to observe changes in L-Arg transport, NOS gene expression and activity and NO generation in the vascular adventitia to determine the mechanism of activation of the L-Arg/NOS/NO pathway. 3. Severe sepsis resulted in severe disturbance of haemodynamic features, with decreased mean arterial blood pressure, brachycardia and inhibited cardiac function (decreased left ventricular +/-dP/dt(max)). Left ventricular end-diastolic pressure was elevated threefold (P < 0.01) under anaesthesia. Rats with sepsis showed severe glucopenia and lacticaemia. Plasma levels of the inflammatory factors macrophage chemoattractant protein-1 and interleukin-8 were increased five- and 29-fold, respectively (P < 0.01). 4. In the adventitia of the thoracic and abdominal aortas, the L-Arg/NO pathway was similarly characterized: the uptake of [(3)H]-L-Arg was Na(+) independent, with the peak occurring at approximately 40 min incubation. Total NOS activity was largely calcium independent (> 90%). The V(max) of L-Arg transport in the sepsis group was increased by 83.5% (P < 0.01), but the K(m) value was not significantly different compared with controls. 5. The mRNA levels of cationic amino acid transporter (CAT)-1 and CAT-2B in the sepsis group were increased by 86 and 62%, respectively (both P < 0.01). Inducible NOS activity was increased 2.8-fold compared with controls (P < 0.01) and iNOS mRNA levels were elevated approximately sixfold (P < 0.01). The NO levels in the plasma and incubation media (incubation for 40 min) in the sepsis group were increased by 144 and 273%, respectively (both P < 0.01). 6. The Arg/NOS/NO pathway was activated in the vascular adventitia of rats with sepsis shock. The L-Arg/NOS/NO pathway in the aortic adventitia may play an important role in the pathogenesis of sepsis and septic shock.