The effects of osmotic stimulation and water availability on c-Fos and FosB staining in the supraoptic and paraventricular nuclei of the hypothalamus.

We studied the effects of osmotic stimulation on the expression of FosB and c-Fos in the supraoptic nucleus (SON) and paraventricular nucleus (PVN). Adult male rats were divided into two groups that were injected with lidocaine (0.1-0.2 ml sc) followed by either 0.9% or 6% NaCl (1 ml/100 g bw sc). After the NaCl injections, the rats were anesthetized and perfused 2, 6, or 8 h after the injections. Their brains were prepared for immunocytochemistry and stained with FosB and c-Fos antibodies. The number of c-Fos-positive cells was significantly increased only at 2 h in the SON and PVN. In contrast, the number of FosB-positive cells was significantly increased at 6, and 8 h in both the SON and PVN. In a second experiment, the effect of water availability on FosB staining 8 h after injections of 6% NaCl was tested in 3 groups of rats: water ad libitum, rats that had no access to water, and rats that were given water 2 h prior to perfusion. FosB staining was significantly reduced in both the SON and the PVN of rats that had ad libitum water compared to the two water-restricted groups. In the third experiment, rats were injected with either 0.9% NaCl or 6% NaCl and were either given ad libitum access to water or water restricted for 6 h after the injections and perfused 24 h after the saline injections. FosB staining was not increased when water was available ad libitum. FosB staining was significantly increased at 24 h in the rats injected with 6% NaCl when water was restricted. Thus, FosB may continue to influence protein expression in the SON and PVN for at least 24 h following acute osmotic stimulation.

FosB expression in the central nervous system following isotonic volume expansion in unanesthetized rats.

We examined c-Fos and FosB staining in the central nervous system 8 and 24 h following acute volume expansion in unanesthetized rats. Male rats were instrumented with a femoral artery catheter for measurement of blood pressure and heart rate (HR), a jugular venous catheter for measurement of central venous pressure (CVP), and a femoral vein catheter for i.v. infusion. After 48 h, rats were volume expanded with isotonic saline (10% of body weight for 10 min i.v.) or given a control infusion (0.01 ml/min for 10 min i.v.). After a period of 8 or 24 h, the rats were deeply anesthetized and perfused transcardially with 4% paraformaldehyde. Separate sets of serial sections of the hypothalamus were processed for either FosB (Santa Cruz) or c-Fos (Oncogene AB-5) immunocytochemistry. The volume expansion protocol significantly increased central venous pressure but did not affect blood pressure or heart rate. Volume expansion produced a significant increase in FosB-positive cells in the paraventricular nucleus (PVN) of the hypothalamus, the supraoptic nucleus (SON), the perinuclear zone (PNZ) of the supraoptic nucleus, the nucleus of the solitary tract (NST), and the caudal ventrolateral medulla (CVL) in both the 8- and 24-h groups. In the area postrema (AP), the number of FosB-positive cells was significantly increased only at 8 h post-infusion. However, c-Fos was not significantly increased above control levels at either time point. The results demonstrate that FosB activation is maintained for at least 24 h following an acute increase in central venous pressure.

Rats exhibit aldosterone-dependent sodium appetite during 24 h hindlimb unloading.

Hindlimb unloading (HU) is an animal model of microgravity and bed rest. In these studies, we examined the role of ingestive behaviours in regulating body fluid balance during 24 h HU. In the first experiment, all rats were given distilled water to drink while two groups were also given access to a sodium chloride solution (0.9% or 1.8%). Water and saline intakes were measured before, during and after 24 h of HU. Rats reduced water intake during 24 h HU in all conditions. During HU, rats increased their intakes of both saline solutions (0.9% NaCl (n= 11): control 7.8 +/- 3 ml; HU 18.2 +/- 4 ml; recovery 8.9 +/- 2.5 ml; 1.8% NaCl (n= 7): control 1.0 +/- 0.4 ml; HU 3.8 +/- 0.3 ml; recovery 1.2 +/- 0.5 ml). Although water intake decreased there was no reduction in total fluid intake when saline was available. Plasma volumes were reduced during HU compared to rats in a normal posture when only water was available to drink (control (n= 11) versus HU (n= 11): 4.0 +/- 0.2 versus 3.4 +/- 0.2 ml (100 g body weight)(-1)). When 0.9% saline was available in addition to water, plasma volumes after 24 h HU were not different from rats in a normal posture (control (n= 11) versus HU (n= 12): 4.3 +/- 0.4 versus 4.3 +/- 0.1 ml (100 g body weight)(-1)). Plasma aldosterone but not plasma renin activity was significantly elevated after 24 h HU. Central infusions of spironolactone blocked the increased intake of 1.8% saline that was associated with 24 h HU. Thus, HU results in an aldosterone-dependent sodium appetite and the ingestion of sodium may help maintain plasma volume.

Cardiovascular regulation of supraoptic vasopressin neurons.

A number of laboratories have identified several key areas in the central nervous system that relay information from arterial baroreceptors to the supraoptic nucleus. Two of these regions are the diagonal band of Broca and the perinuclear zone of the supraoptic nucleus. Recent findings suggest that the inhibition of vasopressin neurons in the SON by caval-atrial stretch may also involve the perinuclear zone. Using Fos immunocytochemistry in combination with volume expansion in unanesthetized rats, we observed that volume expansion activates a number of regions in the CNS including the area postrema, the nucleus of the solitary tract, the caudal ventrolateral medulla, the paraventricular nucleus, the perinuclear zone and oxytocin neurons in the supraoptic nucleus. Further experiments using pericardial catheters demonstrate that the activation of the nucleus of the solitary tract, the ventrolateral medulla, the paraventricular nucleus and the perinuclear zone by volume expansion is dependent on cardiac afferents. However, the Fos in the area postrema and oxytocin neurons of the supraoptic nucleus is not affected by removal of cardiac afferents. Similarly, electrophysiological experiments show that stimulation of cardiac receptors in the caval-atrial junction inhibits supraoptic vasopressin neurons but does not significantly affect the activity of supraoptic oxytocin neurons. These experiments suggest that while the inhibition of supraoptic vasopressin neurons during volume expansion is mediated by cardiac afferents, the activation of supraoptic oxytocin is independent of cardiac afferents and may be mediated by other visceral afferents or humoral factors. Additional electrophysiological experiments examined the importance of the perinuclear zone in cardiopulmonary regulation of vasopressin. Excitotoxin lesions of the perinuclear zone region block the inhibitory effects of caval-atrial stretch on supraoptic vasopressin neurons. This lesion has previously been shown to block the inhibitory effects of arterial baroreceptor stimulation on supraoptic vasopressin neurons. Thus, the neural pathways that inhibit vasopressin release in response to an increase in blood pressure and an increase in blood volume may overlap at the perinuclear zone of the supraoptic nucleus. Also while the inhibition of supraoptic vasopressin neurons during volume expansion is mediated by cardiac afferents, the activation of supraoptic oxytocin neurons is independent of cardiac afferents and may be mediated by other visceral afferents or hormonal factors.

Intrapericardial procaine affects volume expansion-induced fos immunoreactivity in unanesthetized rats.

Acute volume expansion is associated with a specific pattern of Fos expression and the goal of the present study was to evaluate the contribution of cardiac receptors to this response. Adult male rats were instrumented with pericardial catheters introduced at the level of the thymus. Rats were also catheterized for measuring blood pressure, heart rate, central venous pressure, and intravenous infusion. Each rat received a 200-microl intrapericardial (i.p.c) injection of 2% procaine or 0.9% NaCl. Rats were then volume expanded with isotonic saline (10% body weight in 10 min) or given a control infusion (0.01 ml/min for 10 min). Ninety minutes after the start of the infusion, the rats were anesthetized and perfused transcardially. Their brains were sectioned and processed for Fos, dopamine-beta-hydroxylase, and oxytocin immunocytochemistry. Volume expansion plus i.p.c. saline produced a significant increase in Fos expression in the nucleus of the solitary tract, the ventrolateral medulla, the area postrema, the locus coeruleus, the paraventricular nucleus of the hypothalamus, the perinuclear zone of the supraoptic nucleus, and oxytocin neurons in the supraoptic nucleus. The i.p.c. procaine significantly blocked Fos expression produced by the volume expansion in the all of the regions examined except for the area postrema and the SON oxytocin neurons.