Brain volume regulation in response to changes in osmolality.

Hypoosmolality and hyperosmolality are relatively common clinical problems. Many different factors contribute to the substantial morbidity and mortality known to occur during states of altered osmotic homeostasis. The brain is particularly vulnerable to disturbances of body fluid osmolality. The most serious complications are associated with pathological changes in brain volume: brain edema during hypoosmolar states and brain dehydration during hyperosmolar states. Studies in animals have elucidated many of the mechanisms involved with brain adaptation to osmotic stresses, and indicate that it is a complex process involving transient changes in water content and sustained changes in electrolyte and organic osmolyte contents. Appreciation of the nature of the adaptation process, and conversely the deadaptation processes that occur after recovery from hypoosmolality and hyperosmolality, enables a better understanding of the marked variations in neurological sequelae that characterize hyperosmolar and hypoosmolar states, and provides a basis for more rational therapies.

Changes in plasma arginine vasopressin concentrations in cyclists participating in a 109-km cycle race.

OBJECTIVE: To evaluate the osmotic and non-osmotic regulation of arginine vasopressin (AVP) during endurance cycling. DESIGN: Observational study. Setting 109 km cycle race. PARTICIPANTS: 33 Cyclists. INTERVENTIONS: None. MAIN OUTCOME MEASUREMENTS: Plasma sodium concentration ([Na(+)]), plasma volume (PV) and plasma arginine vasopressin (AVP) concentration ([AVP](p)). RESULTS: A fourfold increase in [AVP](p) occurred despite a 2-mmol l(-1) decrease in plasma [Na(+)] combined with only modest (5%) PV contraction. A significant inverse correlation was noted between [AVP](p) Delta and urine osmolality Delta (r = -0.41, p<0.05), whereas non-significant inverse correlations were noted between [AVP](p) and both plasma [Na(+)] Delta and % PV Delta. Four cyclists finished the race with asymptomatic hyponatraemia. The only significant difference between the entire cohort with this subset of athletes was postrace plasma [Na(+)] (137.7 vs 133.5 mmol l(-1), p<0.001) and plasma [Na(+)] Delta (-1.9 vs -5.1 mmol l(-1), p<0.05). The mean prerace [AVP](p) of these four cyclists was just below the minimum detectable limit (0.3 pg ml(-1)) and increased marginally (0.4 pg ml(-1)) despite the decline in plasma [Na(+)]. CONCLUSIONS: The osmotic regulation of [AVP](p) during competitive cycling was overshadowed by non-osmotic AVP secretion. The modest decrease in PV was not the primary non-osmotic stimulus to AVP. Partial suppression of AVP occurred in four (12%) cyclists who developed hyponatraemia during 5 h of riding. Therefore, these results confirm that non-osmotic AVP secretion and exercise-associated hyponatraemia does, in fact, occur in cyclists participating in a 109 km cycle race. However, the stimuli to AVP is likely different between cycling and running.

Acute changes in arginine vasopressin, sweat, urine and serum sodium concentrations in exercising humans: does a coordinated homeostatic relationship exist?

The parallel response of sweat rate and urine production to changes in plasma osmolality and volume support a role for arginine vasopressin (AVP) as the main endocrine regulator of both excretions. A maximal test to exhaustion and a steady-state run on a motorised treadmill were both completed by 10 moderately trained runners, 1 week apart. Sweat, urine and serum sodium concentrations ([Na+]) were evaluated in association with the plasma concentrations of cytokines, neurohypophyseal and natriuretic peptides, and adrenal steroid hormones. When data from both the high-intensity and steady-state runs were combined, significant linear correlations were noted between: sweat [Na+] versus postexercise urine [Na+] (r=0.80; p<0.001), postexercise serum [Na+] versus both postexercise urine [Na+] (r=0.56; p<0.05) and sweat [Na+] (r=0.64; p<0.01) and postexercise urine [Na+] versus postexercise plasma arginine vasopressin concentration ([AVP](P)) (r=0.48; p<0.05). A significant positive correlation was noted between postexercise [AVP](P) and sweat [Na+] during the steady-state condition only (r=0.66; p<0.05). These correlations suggest that changes in serum [Na+] during exercise may evoke corresponding changes in sweat and urine [Na+], which are likely regulated coordinately by changes in [AVP](P) to preserve body fluid homeostasis.

Vasopressin receptor antagonists.

The first non-peptide vasopressin receptor antagonist (VRA) was recently approved by the United States Food and Drug Administration, and several others are now in late stages of clinical development. Phase 3 trials indicate that these agents predictably reduce urine osmolality, increase electrolyte-free water excretion, and raise serum sodium concentration. They are likely to become a mainstay of treatment of euvolemic and hypervolemic hyponatremia. Although tachyphylaxis to the hydro-osmotic effect of these agents does not appear to occur, their use is accompanied by an increase in thirst, and they do not always eliminate altogether the need for water restriction during treatment of hyponatremia. Experience with use of these agents for treatment of acute, severe, life-threatening hyponatremia as well as chronic hyponatremia is limited. Further studies are needed to determine how they are best used in these situations, but the risk of overly rapid correction of hyponatremia seems low. Results of long-term trials to determine the ability of VRAs to reduce morbidity or mortality in congestive heart failure or to slow the progression of polycystic kidney disease are awaited with great interest.

Renal ENaC subunit, Na-K-2Cl and Na-Cl cotransporter abundances in aged, water-restricted F344 x Brown Norway rats.

Renal sodium reabsorption is a key determinant of final urine concentration. Our aim was to determine whether differences existed between aged and young rats in their response to water restriction with regard to the regulation of abundance of any of the major distal renal sodium transporter proteins. Male Fisher 344 x Brown Norway (F344 x BN) rats of 3-, 10-, 24-, or 31 months of age (3M, 10M, 24M, or 31M) were either water restricted (WR) for 5 days or control (ad libitum water). Major renal sodium transporters and channel subunits were evaluated by immunoblotting and immunohistochemistry. Age did not significantly affect plasma arginine vasopressin or aldosterone levels, but renin activity was only 8% in 31M-WR rats relative to 3M-WR (P<0.05). Extreme aging (31M) led to decreased outer medullary abundance of the bumetanide-sensitive Na-K-2Cl cotransporter and decreased cortical abundance of the beta- and gamma-subunits (70-kDa band) of the epithelial sodium channel (ENaC) (P<0.05). Water restriction significantly (P<0.05) increased the abundance of Na-K-2Cl cotransporter (NKCC2) and Na-Cl cotransporter (NCC) across ages. However, these increases were significantly blunted as rats aged. Mean band densities were increased in WR rats (relative to age controls) by 54 and 106% at 3M, but only 25 and 29% at 24M and 0 and 6% at 31M for NKCC2 and NCC, respectively. Aged F344 x BN rats have reduced basal distal tubular renal sodium transporter abundances and blunted upregulation during water restriction, which may contribute to decreased urinary concentrating capacity.

Vasopressin V2 receptor antagonists.

Hyponatremia, whether due to the syndrome of inappropriate antidiuretic hormone secretion (SIADH) or disorders of water retention such as congestive heart failure and cirrhosis, is a very common problem encountered in the care of medical patients. To date, available treatment modalities for disorders of excess arginine vasopressin (AVP) secretion or action have been limited and suboptimal. The recent discovery and development of nonpeptide AVP V(2) receptor antagonists represents a promising new treatment option to directly antagonize the effects of elevated plasma AVP concentrations at the level of the renal collecting ducts. By decreasing the water permeability of renal collecting tubules, excretion of retained water is promoted, thereby normalizing or improving hypo-osmolar hyponatremia. In this review, SIADH and other water retaining disorders are briefly discussed, after which the published preclinical and clinical studies of several nonpeptide AVP V(2) receptor antagonists are summarized. The likely therapeutic indications and potential complications of these compounds are also described.

Glucose effects on gastric motility and tone evoked from the rat dorsal vagal complex.

1. To examine the effects of glucose on the central components of the vago-vagal reflex control of gastric function, we performed both in vivo and in vitro experiments on neurones in the medial nucleus of the tractus solitarius (mNTS) and in the dorsal motor nucleus of the vagus (DMV). 2. In the in vivo anaesthetized rat preparation, unilateral microinjection of D-glucose (10 or 50 mM (60 nl)(-1)) in mNTS produced inhibition of gastric motility and an increase in intragastric pressure. D-glucose had no effect in the DMV. 3. In the in vitro rat brainstem slice preparation, whole-cell recordings of DMV neurones showed that increasing the glucose concentration of the perfusion solution from 5 mM to 15 or 30 mM produced outward currents of 35 +/- 5 pA (n = 7) and 51 +/- 10 pA (n = 11), respectively. These were blocked by tetrodotoxin and picrotoxin, indicating that glucose was acting indirectly to cause the release of GABA. Decreasing the glucose concentration of the perfusing solution by one-half produced an inward current of 36 +/- 5 pA (n = 7). 4. Stimulation of the NTS evoked inhibitory postsynaptic currents (IPSCs) in DMV neurones. The amplitude of the evoked IPSCs was positively correlated with glucose concentration. Perfusion with the ATP-sensitive K(+) (K(ATP)) channel opener diazoxide mimicked the effect of reduced glucose, while perfusion with the K(ATP) channel blocker glibenclamide mimicked the effects of increased glucose. 5. Our data indicate that glucose had no direct excitatory effect on DMV neurones, but DMV neurones appear to be affected by an action of glucose on cell bodies of mNTS neurones via effects on an ATP-sensitive potassium channel.

Increased renal Na-K-ATPase, NCC, and beta-ENaC abundance in obese Zucker rats.

Renal sodium retention, as a result of increased abundance of sodium transporters, may play a role in the development and/or maintenance of the increased blood pressure in obesity. To address this hypothesis, we evaluated the relative abundances of renal sodium transporters in lean and obese Zucker rats at 2 and 4 mo of age by semiquantitative immunoblotting. Mean systolic blood pressure was higher in obese rats relative to lean at 3 mo, P < 0.02. Furthermore, circulating insulin levels were 6- or 13-fold higher in obese rats compared with lean at 2 or 4 mo of age, respectively. The abundances of the alpha(1)-subunit of Na-K-ATPase, the thiazide-sensitive Na-Cl cotransporter (NCC or TSC), and the beta-subunit of the epithelial sodium channel (ENaC) were all significantly increased in the obese rats' kidneys. There were no differences for the sodium hydrogen exchanger (NHE3), the bumetanide-sensitive Na-K-2Cl cotransporter (NKCC2 or BSC1), the type II sodium-phosphate cotransporter (NaPi-2), or the alpha-subunit of ENaC. These selective increases could possibly increase sodium retention by the kidney and therefore could play a role in obesity-related hypertension.

Responses of magnocellular neurons to osmotic stimulation involves coactivation of excitatory and inhibitory input: an experimental and theoretical analysis.

How does a neuron, challenged by an increase in synaptic input, display a response that is independent of the initial level of activity? Here we show that both oxytocin and vasopressin cells in the supraoptic nucleus of normal rats respond to intravenous infusions of hypertonic saline with gradual, linear increases in discharge rate. In hyponatremic rats, oxytocin and vasopressin cells also responded linearly to intravenous infusions of hypertonic saline but with much lower slopes. The linearity of response was surprising, given both the expected nonlinearity of neuronal behavior and the nonlinearity of the oxytocin secretory response to such infusions. We show that a simple computational model can reproduce these responses well, but only if it is assumed that hypertonic infusions coactivate excitatory and inhibitory synaptic inputs. This hypothesis was tested first by applying the GABA(A) antagonist bicuculline to the dendritic zone of the supraoptic nucleus by microdialysis. During local blockade of GABA inputs, the response of oxytocin cells to hypertonic infusion was greatly enhanced. We then went on to directly measure GABA release in the supraoptic nucleus during hypertonic infusion, confirming the predicted rise. Together, the results suggest that hypertonic infusions lead to coactivation of excitatory and inhibitory inputs and that this coactivation may confer appropriate characteristics on the output behavior of oxytocin cells. The nonlinearity of oxytocin secretion that accompanies the linear increase in oxytocin cell firing rate reflects frequency-facilitation of stimulus-secretion coupling at the neurohypophysis.

Vasopressin V2 receptor antagonists.

Hyponatremia due to the syndrome of inappropriate antidiuretic hormone secretion (SIADH) and disorders of water retention such as congestive heart failure and cirrhosis is a common problem encountered in the care of the medical patient. Thus far, available treatment modalities for disorders of excess arginine vasopressin (AVP) secretion or action have been suboptimal. The development of nonpeptide AVP V2 receptor antagonists represents a promising treatment option to directly antagonize the effects of elevated plasma AVP concentrations by increasing the water permeability of renal collecting tubules, thereby promoting excretion of retained water and normalizing hypoosmolar hyponatremia. In this review, SIADH and other water retaining disorders are briefly discussed, after which the published preclinical and clinical studies in the development of several nonpeptide AVP V2 receptor antagonists are summarized. The likely therapeutic indications and potential complications of these compounds, as well as their vascular effects, are also described.

Mineralocorticoid treatment attenuates activation of oxytocinergic and vasopressinergic neurons by icv ANG II.

Central oxytocin (OT) neurons limit intracerebroventricular (icv) ANG II-induced NaCl intake. Because mineralocorticoids synergistically increase ANG II-induced NaCl intake, we hypothesized that mineralocorticoids may attenuate ANG II-induced activation of inhibitory OT neurons. To test this hypothesis, we determined the effect of deoxycorticosterone (DOCA; 2 mg/day) on icv ANG II-induced c-Fos immunoreactivity in OT and vasopressin (VP) neurons in the supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus and also on pituitary OT and VP secretion in male rats. DOCA significantly decreased the percentage of c-Fos-positive (%c-Fos+) OT neurons in the SON and PVN, both in the magnocellular and parvocellular subdivisions, and the %c-Fos+ VP neurons in the SON after a 5-ng icv injection of ANG II. DOCA also significantly reduced the %c-Fos+ OT neurons in the SON after 10 ng ANG II and tended to attenuate 10 ng ANG II-induced OT secretion. However, the %c-Fos+ OT neurons in DOCA-treated rats was greater after 10 ng ANG II, and DOCA did not affect the %c-Fos+ OT neurons in the PVN nor VP secretion or c-Fos immunoreactivity in either the SON or PVN after 10 ng ANG II. DOCA also did not significantly alter the effect of intraperitoneal (ip) cholecystokinin (62 microg) on %c-Fos+ OT neurons or of ip NaCl (2 ml of 2 M NaCl) on the %c-Fos+ OT and VP neurons. These findings indicate that DOCA attenuates the responsiveness of OT and VP neurons to ANG II without completely suppressing the activity of these neurons and, therefore, support the hypothesis that attenuation of OT neuronal activity is one mechanism by which mineralocorticoids enhance NaCl intake.

Chronic hypoosmolality induces a selective decrease in magnocellular neurone soma and nuclear size in the rat hypothalamic supraoptic nucleus.

The magnocellular neurones of the hypothalamo-neurohypophysial system (HNS) play a vital role in the maintenance of body homeostasis by regulating oxytocin (OT) and vasopressin (VP) secretion from the posterior pituitary. During hyperosmolality, OT and VP mRNA levels are known to increase by approximately two-fold, whereas during chronic hypoosmolality, OT and VP mRNA levels decrease to approximately 10-20% of basal levels. In these studies, we evaluated changes in cell size associated with these physiological conditions. Cell and nuclear sizes of neurones in the supraoptic nucleus (SON), the nucleus of the lateral olfactory tract (LOT) and the medial habenular nucleus (MHB) were measured from neurones identified by in situ hybridization histochemistry for beta(III)-tubulin mRNA, and measurements were made from OT and AVP magnocellular neurones in the SON after phenotypic identification by immunohistochemistry. Under hypoosmolar conditions, the cell and nuclear sizes of OT and VP magnocellular neurones decreased to approximately 60% of basal values, whereas cell and nuclear sizes of OT and VP neurones in hyperosmolar rats increased to approximately 170% of basal values. In contrast, neither hyperosmolality, nor hypoosmolality significantly affected cell and nuclear sizes in the LOT and MHB. These results confirm previous studies that showed that magnocellular neurones increase cell size in response to hyperosmolar conditions and, for the first time, demonstrate a marked decrease in cell size in the SON in response to chronic hypoosmolar conditions. These dramatic changes in cell and nuclear size directly parallel changes in OT and VP gene expression in the magnocellular neurones of the SON and, consequently, are consistent with the pronounced bidirectional changes in gene expression and cellular activity found during these osmotic perturbations. Our results therefore support the concept of global alterations in the synthetic activity of magnocellular OT and AVP neurones in response to extracellular osmolality.

Estradiol attenuates angiotensin-induced aldosterone secretion in ovariectomized rats.

Estrogen replacement therapy significantly reduces the risk of cardiovascular disease in postmenopausal women. Previous studies indicate that estradiol (E2) decreases angiotensin II (AT) receptor density in the adrenal and pituitary in NaCl-loaded rats. We used an in vivo model that eliminates the potentially confounding influence of ACTH to determine whether the E2-induced decrease in adrenal AT receptor expression affects aldosterone responses to angiotensin II (Ang II). Female rats were ovariectomized, treated with oil (OVX) or E2 (OVX+E2; 10 microg, s.c.) for 14 days, and fed a NaCl-deficient diet for the last 7 days to maximize adrenal AT receptor expression and responsiveness. On days 12-14 rats were treated with dexamethasone (DEX; 25 microg, i.p., every 12 h) to suppress plasma ACTH. On day 14 aldosterone secretion was measured after a 30-min infusion of Ang II (330 ng/min). Ang II infusion increased the peak plasma aldosterone levels to a lesser degree in the OVX+E2 than in the OVX rats (OVX, 1870 +/- 290 pg/ml; OVX+E2, 1010 +/- 86 pg/ml; P < 0.05). Ang II-induced ACTH and aldosterone secretion was also studied in rats that were not treated with DEX. In the absence of DEX, the peak plasma aldosterone response was also significantly decreased (OVX, 5360 +/- 1200 pg/ml; OVX+E2, 2960 +/- 570 pg/ml; P < 0.05). However, E2 also reduced the plasma ACTH response to Ang II (P < 0.05; OVX, 220 +/- 29 pg/ml; OVX+E2, 160 +/- 20 pg/ml), suggesting that reduced pituitary ACTH responsiveness to Ang II contributes to the effect of E2 on Ang II-induced aldosterone secretion. Adrenal AT1 binding studies confirmed that E2 significantly reduces adrenal AT1 receptor expression in both the presence and absence of DEX in NaCl-deprived rats. These results indicate that E2-induced decreases in pituitary and adrenal AT1 receptor expression are associated with attenuated pituitary ACTH and adrenal aldosterone responses to Ang II and suggest that estrogen replacement therapy may modulate Ang II-stimulated aldosterone secretion as part of its well known cardioprotective actions.

Blood-brain barrier disruption and complement activation in the brain following rapid correction of chronic hyponatremia.

In previous studies we developed a rat model in which demyelination is reproducibly produced following rapid correction of chronic hyponatremia and demonstrated that the development of demyelination in this model is strongly associated with NMR indices of blood-brain barrier (BBB) disruption. Because complement is toxic to oligodendrocytes, we evaluated the hypothesis that BBB disruption precipitated by correction of hypoosmolality is followed by an influx of complement into the brain, which then contributes to the demyelination that occurs under these conditions. We studied four groups of rats with immunocytochemical analysis using primary antibodies to IgG and the C3d split-fragment of activated complement: (1) normal rats; (2) rats in which hyponatremia was maintained for 7 days; (3) chronically hyponatremic rats in which the plasma [Na(+)] was rapidly corrected with hypertonic saline administration 20 h prior to perfusion; and (4) chronically hyponatremic rats in which the plasma [Na(+)] was rapidly corrected with hypertonic saline administration 5 days prior to perfusion. In normonatremic and uncorrected hyponatremic rats only background staining was observed in areas lacking a BBB and in blood vessel walls, whereas marked increases in IgG and C3d staining were seen in the brains of rats both 20 h and 5 days after rapid correction of hyponatremia. The staining intensity was significantly correlated with the degree of neurological impairment. These results provide evidence for functional BBB disruption following rapid correction of hyponatremia and support the hypothesis that complement activation may be involved in the pathogenesis of osmotic demyelination.

Gene expression in the rat supraoptic nucleus induced by chronic hyperosmolality versus hyposmolality.

Magnocellular neurons of the hypothalamo-neurohypophysial system play a fundamental role in the maintenance of body homeostasis by secreting vasopressin and oxytocin in response to systemic osmotic perturbations. During chronic hyperosmolality, vasopressin and oxytocin mRNA levels increase twofold, whereas, during chronic hyposmolality, these mRNA levels decrease to 10-20% of that of normoosmolar control animals. To determine what other genes respond to these osmotic perturbations, we have analyzed gene expression during chronic hyper- versus hyponatremia. Thirty-seven cDNA clones were isolated by differentially screening cDNA libraries that were generated from supraoptic nucleus tissue punches from hyper- or hyponatremic rats. Further analysis of 12 of these cDNAs by in situ hybridization histochemistry confirmed that they are osmotically regulated. These cDNAs represent a variety of functional classes and include cytochrome oxidase, tubulin, Na(+)-K(+)-ATPase, spectrin, PEP-19, calmodulin, GTPase, DnaJ-like, clathrin-associated, synaptic glycoprotein, regulator of GTPase stimulation, and gene for oligodendrocyte lineage-myelin basic proteins. This analysis therefore suggests that adaptation to chronic osmotic stress results in global changes in gene expression in the magnocellular neurons of the supraoptic nucleus.

Vasopressin-mediated regulation of epithelial sodium channel abundance in rat kidney.

Sodium transport is increased by vasopressin in the cortical collecting ducts of rats and rabbits. Here we investigate, by quantitative immunoblotting, the effects of vasopressin on abundances of the epithelial sodium channel (ENaC) subunits (alpha, beta, and gamma) in rat kidney. Seven-day infusion of 1-deamino-[8-D-arginine]-vasopressin (dDAVP) to Brattleboro rats markedly increased whole kidney abundances of beta- and gamma-ENaC (to 238% and 288% of vehicle, respectively), whereas alpha-ENaC was more modestly, yet significantly, increased (to 142% of vehicle). Similarly, 7-day water restriction in Sprague-Dawley rats resulted in significantly increased abundances of beta- and gamma- but no significant change in alpha-ENaC. Acute administration of dDAVP (2 nmol) to Brattleboro rats resulted in modest, but significant, increases in abundance for all ENaC subunits, within 1 h. In conclusion, all three subunits of ENaC are upregulated by vasopressin with temporal and regional differences. These changes are too slow to play a major role in the short-term action of vasopressin to stimulate sodium reabsorption in the collecting duct. Long-term increases in ENaC abundance should add to the short-term regulatory mechanisms (undefined in this study) to enhance sodium transport in the renal collecting duct.

Changes in cerebral blood flow and distribution associated with acute increases in plasma sodium and osmolality of chronic hyponatremic rats.

The cause of the osmotic demyelination syndrome that follows too rapid correction of chronic hyponatremia (CHN) is unknown. Recently, we reported in CHN rats an association between blood-brain barrier (BBB) disruption occurring as early as 3 h into correction and subsequent demyelination. Given the changes in brain water and blood volume which occur during correction of CHN, we hypothesized that the same correction protocol that causes demyelination might alter cerebral blood flow (CBF) during correction, thereby possibly contributing to BBB disruption and demyelination. Ten CHN rats were given hypertonic sodium intraperitoneally and its effect on CBF was continuously monitored for 3 h by magnetic resonance flow imaging. Over the subsequent 3 h, plasma sodium rose from 110.8 to 127.6 mEq/liter (P < 0.001) but neither mean arterial blood pressure nor arterial CO(2) tension changed significantly. By 30 min, CBF increased by 50% in cortical and subcortical areas (P < 0.001) and remained elevated for the next 60 min. After 2 h, cortical flow was no longer elevated significantly and by 3 h it had returned to control values. Subcortical flow, however, significantly exceeded control values throughout the 3 h so that after 2 h the ratio of cortical to subcortical blood flow had fallen from 1.17 to 0.91 (P < 0.05). Although the mechanism by which increased plasma sodium and osmolality alters CBF is uncertain, the results suggest that changes in CBF may be part of a cascade of cerebrovascular disturbances including endothelial or parenchymal damage, mechanical events, metabolic disturbances, or cytokine release which eventually lead to BBB disruption and subsequent demyelination.

Positive association between blood brain barrier disruption and osmotically-induced demyelination.

Rapid correction of chronic hyponatremia can cause osmotic brain demyelination in animals and humans. Why demyelination develops is unknown, but blood brain-barrier disruption might expose oligodendrocytes to substances normally excluded from the brain. To test this hypothesis, chronic hyponatremia was induced and corrected using a new, reproducible rat model for producing osmotic brain demyelination. Blood brain barrier integrity was assessed by NMR imaging at either 3, 16 or 24 h during the first day of correction. Demyelination was determined histopathologically 5 - 6 days later. Of 96 rats studied, demyelination developed 5 - 6 days later in 37 rats, 89% of whom showed barrier disruption. In the 59 rats who did not develop demyelination, 45 (76%) had no barrier disruption. Thus, blood-brain barrier disruption during the first 24 h of correction was associated with a 70% risk of developing demyelination. By contrast, the risk of developing subsequent demyelination was only 8% when the barrier was intact. This strong association between barrier disruption and subsequent demyelination provides new insights into the role of blood brain barrier function in demyelinative disorders such as the osmotic demyelination syndrome and by extension to other demyelinative disorders such as multiple sclerosis.