High-precision K isotopic analysis of cerebrospinal fluid and blood serum microsamples via multicollector inductively coupled plasma-mass spectrometry equipped with 1013 Ω faraday cup amplifier resistors.

BACKGROUND: Potassium isotopic analysis is increasingly performed in both geological and biological contexts as a result of the introduction of MC-ICP-MS instrumentation either equipped with a collision/reaction cell or having the capability of working at "extra-high" mass resolution in order to deal with spectral interference caused by argon hydride (ArH+) ions. Potassium plays an important role in the central nervous system, and its isotopic analysis could provide an enhanced insight into the corresponding processes, but K isotopic analysis of cerebrospinal fluid is challenging due to the small volume, a few microliter only, typically available. This work aimed at developing a method for determining the K isotopic signature of serum and cerebrospinal fluid at a final K concentration of 25 ng mL-1 using Faraday cup amplifiers equipped with a 1013 Ω resistor. RESULTS: Potassium isotope ratios obtained for reference materials measured at a final K concentration of 25 ng mL-1 were in excellent agreement with the corresponding reference values and the internal and external precision for the δ41K value was 0.11 ‰ (2SE, N = 50) and 0.10 ‰ (2SD, N = 6), respectively. The robustness against the presence of matrix elements and the concentration mismatch between sample and standard observed at higher K concentrations is preserved at low K concentration. Finally, K isotopic analysis of serum and cerebrospinal fluid (3-12 µL of sample) of healthy mice of both sexes was performed, revealing a trend towards an isotopically lighter signature for serum and cerebrospinal fluid from female individuals, however being significant for serum only. SIGNIFICANCE: This work provides a robust method for high-precision K isotopic analysis at a concentration of 25 ng mL-1. By monitoring both K isotopes, 39K and 41K, with Faraday cups connected to amplifiers with 1013 Ω resistors, accurate K isotope ratio results are obtained with a two-fold improvement in internal and external precision compared to those obtained with the set-up with traditional 1011 Ω resistors. The difference in the K isotope ratio in CSF and serum between the sexes, is possibly indicating an influence of the sex or hormones on the fractionation effects accompanying cellular uptake/release.

Hydrocephalus in a rat model of Meckel Gruber syndrome with a TMEM67 mutation.

Transmembrane protein 67 (TMEM67) is mutated in Meckel Gruber Syndrome type 3 (MKS3) resulting in a pleiotropic phenotype with hydrocephalus and renal cystic disease in both humans and rodent models. The precise pathogenic mechanisms remain undetermined. Herein it is reported for the first time that a point mutation of TMEM67 leads to a gene dose-dependent hydrocephalic phenotype in the Wistar polycystic kidney (Wpk) rat. Animals with TMEM67 heterozygous mutations manifest slowly progressing hydrocephalus, observed during the postnatal period and continuing into adulthood. These animals have no overt renal phenotype. The TMEM67 homozygous mutant rats have severe ventriculomegaly as well as severe polycystic kidney disease and die during the neonatal period. Protein localization in choroid plexus epithelial cells indicates that aquaporin 1 and claudin-1 both remain normally polarized in all genotypes. The choroid plexus epithelial cells may have selectively enhanced permeability as evidenced by increased Na+, K+ and Cl- in the cerebrospinal fluid of the severely hydrocephalic animals. Collectively, these results suggest that TMEM67 is required for the regulation of choroid plexus epithelial cell fluid and electrolyte homeostasis. The Wpk rat model, orthologous to human MKS3, provides a unique platform to study the development of both severe and mild hydrocephalus.

Differences in Sampling Site on Postmortem Cerebrospinal Fluid Biochemistry: A Preliminary Study.

Cerebrospinal fluid (CSF) is often analyzed at postmortem. The presented preliminary study compared postmortem CSF samples for biochemical analysis from the subarachnoid space around the spinal cord and ventricular space of the brain. This study compared 15 paired CSF samples in which the CSF from the subarachnoid space via lumbar puncture had higher sodium and chloride levels and lower magnesium and potassium levels than CSF from the ventricles. The differences correlated significantly with the deceased's age and had a similar trend with postmortem interval. This study suggests that CSF from different collection sites has different electrolyte concentrations, which are age and possibly postmortem interval dependent. When collecting CSF, the pathologist should document the collection site, age, and postmortem interval, and the mixing of CSF samples from different sites should be avoided. Further studies are warranted to clarify other possible reasons to explain the observed differences.

Chronic high-sodium diet intake after weaning lead to neurogenic hypertension in adult Wistar rats.

In this study, we investigated some mechanisms involved in sodium-dependent hypertension of rats exposed to chronic salt (NaCl) intake from weaning until adult age. Weaned male Wistar rats were placed under high (0.90% w/w, HS) or regular (0.27% w/w, Cont) sodium diets for 12 weeks. Water consumption, urine output and sodium excretion were higher in HS rats compared to control. Blood pressure (BP) was directly measured by the arterial catheter and found 13.8% higher in HS vs Cont rats. Ganglionic blockade with hexamethonium caused greater fall in the BP of HS rats (33%), and central antagonism of AT1 receptors (losartan) microinjected into the lateral ventricle reduced BP level of HS, but not of Cont group. Heart rate variability analysis revealed sympathetic prevalence on modulation of the systolic interval. HS diet did not affect creatinine clearance. Kidney histological analysis revealed no significant change in renal corpuscle structure. Sodium and potassium concentrations in CSF were found higher in HS rats despite no change in plasma concentration of these ions. Taken together, data suggest that animals exposed to chronic salt intake to a level close to that reported for human' diet since weaning lead to hypertension, which appears to rely on sodium-driven neurogenic mechanisms.

Disrupted hippocampal network physiology following PTEN deletion from newborn dentate granule cells.

Abnormal hippocampal granule cells are present in patients with temporal lobe epilepsy, and are a prominent feature of most animal models of the disease. These abnormal cells are hypothesized to contribute to epileptogenesis. Isolating the specific effects of abnormal granule cells on hippocampal physiology, however, has been difficult in traditional temporal lobe epilepsy models. While epilepsy induction in these models consistently produces abnormal granule cells, the causative insults also induce widespread cell death among hippocampal, cortical and subcortical structures. Recently, we demonstrated that introducing morphologically abnormal granule cells into an otherwise normal mouse brain - by selectively deleting the mTOR pathway inhibitor PTEN from postnatally-generated granule cells - produced hippocampal and cortical seizures. Here, we conducted acute slice field potential recordings to assess the impact of these cells on hippocampal function. PTEN deletion from a subset of granule cells reproduced aberrant responses present in traditional epilepsy models, including enhanced excitatory post-synaptic potentials (fEPSPs) and multiple, rather than single, population spikes in response to perforant path stimulation. These findings provide new evidence that abnormal granule cells initiate a process of epileptogenesis - in the absence of widespread cell death - which culminates in an abnormal dentate network similar to other models of temporal lobe epilepsy. Findings are consistent with the hypothesis that accumulation of abnormal granule cells is a common mechanism of temporal lobe epileptogenesis.

Analysis of serum and cerebrospinal fluid in clinically normal adult miniature donkeys.

AIM: To establish reference intervals for serum and cerebrospinal fluid (CSF) parameters in clinically healthy adult miniature donkeys. METHODS: Experiments were conducted on 10 female and 10 male clinically normal adult miniature donkeys, randomly selected from five herds. Lumbosacral CSF collection was performed with the sedated donkey in the standing position. Cell analysis was performed immediately after the samples were collected. Blood samples were obtained from the jugular vein immediately after CSF sample collection. Sodium, potassium, glucose, urea nitrogen, total protein, calcium, chloride, phosphorous and magnesium concentrations were measured in CSF and serum samples. A paired t-test was used to compare mean values between female and male donkeys. RESULTS: The CSF was uniformly clear, colourless and free from flocculent material, with a specific gravity of 1.002. The range of total nucleated cell counts was 2-4 cells/µL. The differential white cell count comprised only small lymphocytes. No erythrocytes or polymorphonuclear cells were observed on cytological examination. Reference values were obtained for biochemical analysis of serum and CSF. Gender had no effect on any variables measured in serum or CSF (p>0.05). CONCLUSION AND CLINICAL RELEVANCE: CSF analysis can provide important information in addition to that gained by clinical examination. CSF analysis has not previously been performed in miniature donkeys; this is the first report on the subject. In the present study, reference intervals for total nucleated cell count, total protein, glucose, urea nitrogen, sodium, potassium, chloride, calcium, phosphorous and magnesium concentrations of serum and CSF were determined for male and female miniature donkeys.

Na+ and K+ ion imbalances in Alzheimer's disease.

Alzheimer's disease (AD) is associated with impaired glutamate clearance and depressed Na(+)/K(+) ATPase levels in AD brain that might lead to a cellular ion imbalance. To test this hypothesis, [Na(+)] and [K(+)] were analyzed in postmortem brain samples of 12 normal and 16 AD individuals, and in cerebrospinal fluid (CSF) from AD patients and matched controls. Statistically significant increases in [Na(+)] in frontal (25%) and parietal cortex (20%) and in cerebellar [K(+)] (15%) were observed in AD samples compared to controls. CSF from AD patients and matched controls exhibited no differences, suggesting that tissue ion imbalances reflected changes in the intracellular compartment. Differences in cation concentrations between normal and AD brain samples were modeled by a 2-fold increase in intracellular [Na(+)] and an 8-15% increase in intracellular [K(+)]. Since amyloid beta peptide (Aß) is an important contributor to AD brain pathology, we assessed how Aß affects ion homeostasis in primary murine astrocytes, the most abundant cells in brain tissue. We demonstrate that treatment of astrocytes with the Aß 25-35 peptide increases intracellular levels of Na(+) (~2-3-fold) and K(+) (~1.5-fold), which were associated with reduced levels of Na(+)/K(+) ATPase and the Na(+)-dependent glutamate transporters, GLAST and GLT-1. Similar increases in astrocytic Na(+) and K(+) levels were also caused by Aß 1-40, but not by Aß 1-42 treatment. Our study suggests a previously unrecognized impairment in AD brain cell ion homeostasis that might be triggered by Aß and could significantly affect electrophysiological activity of brain cells, contributing to the pathophysiology of AD.

Investigating paradoxical hysteresis effects in the mouse neocortical slice model.

Clinically, anesthetic drugs show hysteresis in the plasma drug concentrations at induction versus emergence from anesthesia induced unconsciousness. This is assumed to be the result of pharmacokinetic lag between the plasma and brain effect-site and vice versa. However, recent mathematical and experimental studies demonstrate that anesthetic hysteresis might be due in part to lag in the brain physiology, independent of drug transport delay - so-called "neural inertia". The aim of this study was to investigate neural inertia in the reduced neocortical mouse slice model. Seizure-like event (SLE) activity was generated by exposing cortical slices to no-magnesium artificial cerebrospinal fluid (aCSF). Concentration-effect loops were generated by manipulating SLE frequency, using the general anesthetic drug etomidate and by altering the aCSF magnesium concentration. The etomidate (24 µM) concentration-effect relationship showed a clear hysteresis, consistent with the slow diffusion of etomidate into slice tissue. Manipulation of tissue excitability, using either carbachol (50 µM) or elevated potassium (5mM vs 2.5mM) did not significantly alter the size of etomidate hysteresis loops. Hysteresis in the magnesium concentration-effect relationship was evident, but only when the starting condition was magnesium-containing "normal" aCSF. The in vitro cortical slice manifests pathway-dependent "neural inertia" and may be a valuable model for future investigations into the mechanisms of neural inertia in the cerebral cortex.

Capillary endothelial Na(+), K(+), ATPase transporter homeostasis and a new theory for migraine pathophysiology.

BACKGROUND: Cerebrospinal fluid sodium concentration ([Na(+)](csf)) increases during migraine, but the cause of the increase is not known. OBJECTIVE: Analyze biochemical pathways that influence [Na(+)](csf) to identify mechanisms that are consistent with migraine. METHOD: We reviewed sodium physiology and biochemistry publications for links to migraine and pain. RESULTS: Increased capillary endothelial cell (CEC) Na(+), K(+), -ATPase transporter (NKAT) activity is probably the primary cause of increased [Na(+)](csf). Physiological fluctuations of all NKAT regulators in blood, many known to be involved in migraine, are monitored by receptors on the luminal wall of brain CECs; signals are then transduced to their abluminal NKATs that alter brain extracellular sodium ([Na(+)](e)) and potassium ([K(+)](e)). CONCLUSIONS: We propose a theoretical mechanism for aura and migraine when NKAT activity shifts outside normal limits: (1) CEC NKAT activity below a lower limit increases [K(+)](e), facilitates cortical spreading depression, and causes aura; (2) CEC NKAT activity above an upper limit elevates [Na(+)](e), increases neuronal excitability, and causes migraine; (3) migraine-without-aura may arise from CEC NKAT over-activity without requiring a prior decrease in activity and its consequent spreading depression; (4) migraine triggers disturb, and treatments improve, CEC NKAT homeostasis; (5) CEC NKAT-induced regulation of neural and vasomotor excitability coordinates vascular and neuronal activities, and includes occasional pathology from CEC NKAT-induced apoptosis or cerebral infarction.

Neuropeptides modulate compound postsynaptic potentials in basolateral amygdala.

Previous behavioral studies have shown that neuropeptides intrinsic to the amygdala formation can alter fear and anxiety states. We have previously shown that the anxiogenic neuropeptide cholecystokinin (CCK) increases inhibitory neurotransmission in basolateral amygdala. We have since observed that CCK induces synchronized rhythmic activity composed of compound postsynaptic potentials (cPSPs). We have now further characterized these cPSPs by inducing cPSPs routinely in 5 mM extracellular K(+). CCK facilitated cPSP occurrence in a dose dependent manner in brain slices from both young and mature rats. The cPSPs were attenuated by glutamate receptor antagonists (NBQX or DL-AP5) or low concentrations of GABA(A) receptor antagonists (bicuculline methiodide (BMI), SR95531, or picrotoxin), but not by the GABA(B) receptor antagonist, CGP52432. Low concentrations of tetrodotoxin (TTX, 10 nM) also attenuated the cPSPs. The Na-K-2Cl cotransporter blocker, bumetanide (1 or 10 microM) also blocked the cPSPs. The anxiogenic neuropeptide corticotropin-releasing factor (CRF) facilitated cPSPs while anxiolytic neuropeptides (neuropeptide Y (NPY) and somatostatin) attenuated cPSPs. The benzodiazepine agonist diazepam dose-dependently modulated cPSPs. Mefloquine facilitated cPSPs within 10 min of application. We hypothesize that cPSPs are generated by positive feedback between a subset of interneurons and a subset of glutamatergic projection neurons.

Increases in spinal cerebrospinal fluid potassium concentration do not increase isoflurane minimum alveolar concentration in rats.

BACKGROUND: Previous studies demonstrated that MAC for isoflurane directly correlates with the concentration of Na(+) in cerebrospinal fluid surrounding the spinal cord, the primary site for mediation of the immobility produced by inhaled anesthetics. If this correlation resulted from increased irritability of the cord, then infusion of increased concentrations of potassium (K(+)) might be predicted to act similarly. However, an absence of effect of K(+) might be interpreted to indicate that K(+) channels do not mediate the immobility produced by inhaled anesthetics whereas Na(+) channels remain as potential mediators. Accordingly, in the present study, we examined the effect of altering intrathecal concentrations of K(+) on MAC. METHODS: In rats prepared with chronic indwelling intrathecal catheters, we infused solutions deficient in K(+) and with an excess of K(+) into the lumbar space and measured MAC for isoflurane 24 h before, during, and 24 h after infusion. Rats similarly prepared were tested for the effect of altered osmolarity on MAC (accomplished by infusion of mannitol) and for the penetration of Na(+) into the cord. RESULTS: MAC of isoflurane never significantly increased with increasing concentrations of K(+) infused intrathecally. At infused concentrations exceeding 12 times the normal concentration of KCl, i.e., 29 mEq/L, rats moved spontaneously at isoflurane concentrations just below, and sometimes at MAC, but the average MAC in these rats did not exceed their control MAC. At the largest infused concentration (58.1 mEq/L), MAC significantly decreased and did not subsequently return to normal (i.e., such large concentrations produced injury). Infusions of lower concentrations of K(+) had no effect on MAC. Infusion of osmotically equivalent solutions of mannitol did not affect MAC. Na(+) infused intrathecally measurably penetrated the spinal cord. CONCLUSIONS: The results do not support a mediation or modulation of MAC by K(+) channels.

Estimation of time since death from CSF electrolyte concentration in Bhopal region of central India.

Estimation of time since death from sodium and potassium ion concentration levels in CSF (cerebrospinal fluid) was carried out in 100 medico legal autopsies with known time of death in the department of Forensic medicine, Gandhi Medical College in Bhopal region of Central India. CSF was aspirated from lateral ventricles after opening the skull and dura, and concentration of these ions were estimated by flame photometry. Results revealed a significant correlation of sodium and potassium ions in CSF up to 25 h of time since death, with average per hour rise of 1.21 meq/h for potassium and fall of 1.115 meq/h for sodium ions. A useful relationship between sodium potassium ion ratio and PMI (post-mortem interval) was also elicited. The study concludes that changes in CSF electrolyte is a significant parameter to estimate time since death.

Cerebrospinal fluid sodium increases in migraine.

BACKGROUND: Pharmaceuticals with calcium- or sodium-channel-blocking activity have proven useful for migraine prophylaxis, and calcium channel, sodium transporter, and sodium channel gene mutations have been found in familial hemiplegic migraine. However, it is not known whether calcium or sodium homeostasis is altered in migraine. OBJECTIVE: To compare levels of sodium, calcium, potassium, and magnesium in cerebrospinal fluid (CSF) and blood plasma between migraineurs and controls. METHODS: We recruited 20 migraineurs without aura and 11 controls prospectively, and studied migraineurs in sick (MH(+)) and well (MH(-)) states. We collected lumbar CSF and venous blood plasma, quantified elements with ion-selective electrodes or colorimetry, and determined osmolality by depression of freezing point. We compared levels of Na(+), Ca(2+), K(+), and Mg among and also within subjects who were studied in both MH(+) and MH(-) states. RESULTS: Mean CSF Na(+) levels were increased by 3 mmol/L in MH(+) compared with MH(-) and by 4 mmol/L compared to controls (P < 0.005). In 4 subjects who were sampled in both MH(+) and MH(-) states, mean CSF Na(+) concentration increased by 2 mmol/L in the MH(+) state compared with the MH(-) state (P < 0.05). Simultaneous plasma Na(+) levels did not differ among the 3 clinical groups, nor did osmolality, total Ca and Ca(2+), K(+), and total Mg levels in CSF. CONCLUSIONS: Compared to both controls and the MH(-) state, CSF Na(+) concentration increased in MH(+) independently from other clinical or pharmacological fluctuations, CSF concentrations of Ca(2+), Mg, and K(+), and blood plasma Na(+) levels. These results implicate a deviation of Na(+) homeostasis in migraine. The modestly elevated extracellular Na(+) in MH(+) may cause the neural changes that underlie clinical features of migraine.

Effects of potassium concentration on firing patterns of low-calcium epileptiform activity in anesthetized rat hippocampus: inducing of persistent spike activity.

PURPOSE: It has been shown that a low-calcium high-potassium solution can generate ictal-like epileptiform activity in vitro and in vivo. Moreover, during status epileptiform activity, the concentration of [K+]o increases, and the concentration of [Ca2+]o decreases in brain tissue. Therefore we tested the hypothesis that long-lasting persistent spike activity, similar to one of the patterns of status epilepticus, could be generated by a high-potassium, low-calcium solution in the hippocampus in vivo. METHODS: Artificial cerebrospinal fluid was perfused over the surface of the exposed left dorsal hippocampus of anesthetized rats. A stimulating electrode and a recording probe were placed in the CA1 region. RESULTS: By elevating K+ concentration from 6 to 12 mM in the perfusate solution, the typical firing pattern of low-calcium ictal bursts was transformed into persistent spike activity in the CA1 region with synaptic transmission being suppressed by calcium chelator EGTA. The activity was characterized by double spikes repeated at a frequency approximately 4 Hz that could last for >1 h. The analysis of multiple unit activity showed that both elevating [K+]o and lowering [Ca2+]o decreased the inhibition period after the response of paired-pulse stimulation, indicating a suppression of the after-hyperpolarization (AHP) activity. CONCLUSIONS: These results suggest that persistent status epilepticus-like spike activity can be induced by nonsynaptic mechanisms when synaptic transmission is blocked. The unique double-spike pattern of this activity is presumably caused by higher K+ concentration augmenting the frequency of typical low-calcium nonsynaptic burst activity.

Activation of brain renin-angiotensin-aldosterone system by central sodium in Wistar rats.

Functional studies indicate that the sympathoexcitatory and pressor responses to an increase in cerebrospinal fluid (CSF) [Na+] by central infusion of Na+-rich artificial cerebrospinal fluid (aCSF) in Wistar rats are mediated in the brain by mineralocorticoid receptor (MR) activation, ouabain-like compounds (OLC), and AT1-receptor stimulation. In the present study, we examined whether increasing CSF [Na+] by intracerebroventricular infusion of Na+-rich aCSF activates MR and thereby increases OLC and components of the renin-angiotensin system in the brain. Male Wistar rats received via osmotic minipump an intracerebroventricular infusion of aCSF or Na+-rich aCSF, in some groups combined with intracerebroventricular infusion of spironolactone (100 ng/h), antibody Fab fragments (to bind OLC), or as control gamma-globulins. After 2 wk of infusion, resting blood pressure and heart rate were recorded, OLC and aldosterone content in the hypothalamus were assessed by a specific ELISA or radioimmunoassay, and angiotensin-converting enzyme (ACE) and AT1-receptor binding densities in various brain nuclei were measured by autoradiography using 125I-labeled 351 A and 125I-labeled ANG II. When compared with intracerebroventricular aCSF, intracerebroventricular Na+-rich aCSF increased CSF [Na+] by approximately 5 mmol/l, mean arterial pressure by approximately 20 mmHg, heart rate by approximately 65 beats/min, and hypothalamic content of OLC by 50% and of aldosterone by 33%. Intracerebroventricular spironolactone did not affect CSF [Na+] but blocked the Na+-rich aCSF-induced increases in blood pressure and heart rate and OLC content. Intracerebroventricular Na+-rich aCSF increased ACE and AT1-receptor-binding densities in several brain nuclei, and Fab fragments blocked these increases. These data indicate that in Wistar rats, a chronic increase in CSF [Na+] may increase hypothalamic aldosterone and activate CNS pathways involving MR, and OLC, leading to increases in AT1-receptor and ACE densities in brain areas involved in cardiovascular regulation and hypertension.

Cerebrospinal fluid: postmortem biochemical study.

UNLABELLED: Postmortem phenomena can change and alter biochemical components in body fluids such as blood and as cerebrospinal fluid (CSF). AIMS OF THE STUDY WERE: (a) to analyse urea, glucose, potassium, chloride, protein, creatinine, calcium, alkaline phosphatase and cortisol in CSF fluid and (b) to compare results between two age groups, between groups with or without mental or degenerative neurological illness and between a group reported as dying from natural causes and a group that had a violent death. MATERIALS AND METHODS: The study was carried out in Hospitalet de Llobregat (Barcelona) of 55 corpses. Samples were obtained following section of the corpus callosus, through the lateral ventricles and frozen to -80 degrees C until processed. RESULTS: Significant differences were found in urea levels between the two age groups, in protein between natural and violent death groups and in alkaline phosphatase between the two age groups and between the natural and violent death group. Cortisol levels revealed significant difference between the two age groups and is those supplying natural and violent death. CONCLUSIONS: The study indicates to the need for further studies designed to include groups with defined diagnose of mental or degenerative disorders as well as different age groups.

Testing of potassium, sodium and chlorine in liquor at the patients with diabetes mellitus.

Patient with diabetes is susceptible to development of vascular lesions of the blood vessels of all sizes. During the discase there is a progressive development of microangiopatys and arterioscleroses. Continuing the works of Stanley I., Rapaport and other rescarches about changes of permeability of hcmatoenchepal barrier in experimental conditions, (according1) the changes of the barrier at patients with diabetes mellitus was examined in this work. At the patients with diabetes mellitus level of sodium in liquor increases, and the level of chlorine and potassium in liquor decreases.

Ischemia triggered by red blood cell products in the subarachnoid space is inhibited by nimodipine administration or moderate volume expansion/hemodilution in rats.

OBJECTIVE: It has been proposed that delayed ischemic neurological deficits are induced by red blood cell (RBC) products after subarachnoid hemorrhage. Prophylactic treatment with the Ca2+ antagonist nimodipine or prevention of systemic volume contraction reduces the occurrence of delayed ischemic neurological deficits. To gain insight into the underlying mechanism, we studied the effects of nimodipine or volume expansion on ischemic events induced by RBC products in rats. METHODS: A cranial window was implanted in 52 rats. At the window, cerebral blood flow (measured with laser Doppler flowmetry) and the subarachnoid direct current potential were recorded; the cortical surface was superfused with artificial cerebrospinal fluid. A spreading neuronal/astroglial depolarization wave was triggered at a remote site, from which it traveled to the cranial window. RESULTS: In 16 rats, the depolarization wave triggered an ischemic event at the cranial window when artificial cerebrospinal fluid containing the RBC product hemoglobin and elevated K+ levels was superfused. In contrast, in animals receiving intravenously administered nimodipine (n = 12) or moderate volume expansion/hemodilution with hydroxyethyl starch (6% hydroxyethyl starch 200/0.5) (n = 10), the depolarization wave triggered brief initial hypoperfusion, followed by brief hyperemia, in the cortical area exposed to the RBC products. Under physiological conditions, the depolarization wave triggered brief hyperemia (n = 14). CONCLUSION: Spreading ischemia induced by RBC products is antagonized by measures known to be beneficial in the prophylaxis of delayed ischemic neurological deficits. Our findings suggest that a mechanism involving the cortical microcirculation might underlie the therapeutic effects of nimodipine and volume expansion.

Confocal imaging of organic anion transport in intact rat choroid plexus.

We used confocal microscopy and quantitative image analysis to follow the movement of the fluorescent organic anion fluorescein (FL) from bath to cell and cell to blood vessel in intact rat lateral choroid plexus. FL accumulation in epithelial cells and underlying vessels was rapid, concentrative, and reduced by other organic anions. At steady state, cell fluorescence exceeded bath fluorescence by a factor of 3-5, and vessel fluorescence exceeded cell fluorescence by a factor of approximately 2. In cells, FL distributed between diffuse and punctate compartments. Cell and vessel accumulation of FL decreased when metabolism was inhibited by KCN, when bath Na(+) was reduced from 130 to 26 mM, and when the Na(+) gradient was collapsed with ouabain. Cell and vessel accumulation increased by >50% when 1-10 microM glutarate was added to the bath. Finally, transport of FL and carboxyfluorescein (generated intracellularly from carboxyfluorescein diacetate) from cell to blood vessel was greatly diminished when medium K(+) concentration ([K(+)]) was increased 10-fold. These results 1) validate a new approach to the study of choroid plexus function, and 2) indicate a two-step mechanism for transepithelial organic anion transport: indirect coupling of uptake to Na(+) at the apical membrane and electrical potential-driven efflux at the basolateral membrane.

Enhanced sympathoexcitatory and pressor responses to central Na+ in Dahl salt-sensitive vs. -resistant rats.

An enhanced responsiveness to increases in cerebrospinal fluid (CSF) Na+ by high salt intake may contribute to salt-sensitive hypertension in Dahl salt-sensitive (S) rats. To test this hypothesis, sympathetic and pressor responses to acute and chronic increases in CSF Na+ were evaluated. In conscious young (5-6 wk old) and adult (10-11 wk old) Dahl S and salt-resistant (R) rats as well as weight-matched Wistar rats, hemodynamic [blood pressure (BP) and heart rate (HR)] and sympathetic [renal sympathetic nerve activity (RSNA)] responses to 10-min intracerebroventricular infusions of artificial CSF (aCSF) and Na+-rich aCSF (containing 0.2-0.45 M Na+) were evaluated. Intracerebroventricular Na+-rich aCSF increased BP, RSNA, and HR in a dose-related manner. The extent of these increases was significantly larger in Dahl S versus Dahl R or Wistar rats and young versus adult Dahl S rats. In a second set of experiments, young Dahl S and R rats received a chronic intracerebroventricular infusion of aCSF or Na+-rich (0.8 M) aCSF (5 microl/h) for 14 days, with the use of osmotic minipumps. On day 14 in conscious rats, CSF was sampled and BP, HR, and RSNA were recorded at rest and in response to air stress, intracerebroventricular alpha2-adrenoceptor agonist guanabenz, intracerebroventricular ouabain, and intravenous phenylephrine and nitroprusside to estimate baroreflex function. The infusion of Na+-rich aCSF versus aCSF increased CSF Na+ concentration to the same extent but caused severe versus mild hypertension in Dahl S and Dahl R rats, respectively. After central Na+ loading, hypothalamus "ouabain" significantly increased in Dahl S and only tended to increase in Dahl R rats. Moreover, sympathoexcitatory and pressor responses to intracerebroventricular exogenous ouabain were attenuated by Na+-rich aCSF to a greater extent in Dahl S versus Dahl R rats. Responses to air-jet stress or intracerebroventricular guanabenz were enhanced by Na+-rich aCSF in both strains, but the extent of enhancement was significantly larger in Dahl S versus Dahl R. Na+-rich aCSF impaired arterial baroreflex control of RSNA more markedly in Dahl S versus R rats. These findings indicate that genetic control of mechanisms linking CSF Na+ with brain "ouabain" is altered in Dahl S rats toward sympathetic hyperactivity and hypertension.