This study investigated the importance of supraspinal vasopressin and glutamate neurones in regulating renal sympathetic activity as part of the response to an acute reduction in blood volume. Wistar rats anaesthetized with chloralose and urethane were instrumented to record arterial blood pressure, heart rate and left renal sympathetic nerve activity. Pharmacological agonists and antagonists to glutamate and vasopressin were applied to the renal outflow of the spinal cord via an intrathecal catheter inserted at the foramen magnum and with the tip at the level of T10. Both glutamate and vasopressin increased renal sympathetic activity, and these actions were shown to be selectively blocked by their respective antagonists. Removing 1 ml of venous blood from a femoral venous catheter elicited an increase of 26 +/- 2% in renal sympathetic activity. This response to mild haemorrhage was halved to 13 +/- 4% by prior intrathecal application of a selective V1a antagonist. Similarly, prior intrathecal application of kynurenic acid reduced the response to the mild haemorrhage from 28 +/- 2 to 12.6 +/- 2.8%. Intrathecal application of both antagonists together reduced the haemorrhage response even further to 8 +/- 3%. All the changes were statistically significant at P <0.01. It is concluded that a small reduction in blood volume induces an increase in renal sympathetic activity dependent on vasopressin and glutamate release from terminals of supraspinal neurones. It is suggested that the vasopressin neurones most probably originate from the paraventricular nucleus of the hypothalamus.