Hyperosmotic Saline Research Articles

The Prodrug DHED Delivers 17β-Estradiol into the Retina for Protection of Retinal Ganglion Cells and Preservation of Visual Function in an Animal Model of Glaucoma.

We report a three-pronged phenotypic evaluation of the bioprecursor prodrug 10β,17β-dihydroxyestra-1,4-dien-3-one (DHED) that selectively produces 17β-estradiol (E2) in the retina after topical administration and halts glaucomatous neurodegeneration in a male rat model of the disease. Ocular hypertension (OHT) was induced by hyperosmotic saline injection into an episcleral vein of the eye. Animals received daily DHED eye drops for 12 weeks. Deterioration of visual acuity and contrast sensitivity by OHT in these animals were markedly prevented by the DHED-derived E2 with concomitant preservation of retinal ganglion cells and their axons. In addition, we utilized targeted retina proteomics and a previously established panel of proteins as preclinical biomarkers in the context of OHT-induced neurodegeneration as a characteristic process of the disease. The prodrug treatment provided retina-targeted remediation against the glaucomatous dysregulations of these surrogate endpoints without increasing circulating E2 levels. Collectively, the demonstrated significant neuroprotective effect by the DHED-derived E2 in the selected animal model of glaucoma supports the translational potential of our presented ocular neuroprotective approach owing to its inherent therapeutic safety and efficacy.

Novel Treatment Options for Corneal Edema

Introduction: The current treatment of corneal edema is limited to hyperosmotic saline mixtures. Usually the therapy consists of treatment with hypertonic 5% NaCl eye drops or 6% eye ointment. However, these only remain on the cornea for a short time because they are quickly removed by the tear film and blinking. This problem is partly addressed by adding hyaluronic acid to some pharmaceuticals. In this work, Sovermol as a carrier oil supplied with osmotically active substances, honey, icing sugar, Substance C188 and Prevor® substance X were investigated in an experimental setting as therapeutics for corneal edema. Material and Methods: In the Ex vivo Eye irritation Test (EVEIT), corneas can be examined under defined and stable biochemical conditions without blinking. Corneal edema can be simulated and examined using ultrasound pachymetry and Optical Coherence Tomography (OCT). Thus, we investigated the deswelling effect of the substances described above on 23 EVEIT corneas. Results: The mixtures, examined in this study, showed a significant reduction of corneal edema especially mixtures of Sovermol with Substance C188 were very promising as a new approach to treatment of corneal edema. As reference substance for corneal edema treatment ODM5® was used. Other mixtures, especially Honey and Prevor® substance X in DOL showed a sufficient reduction of corneal edema, while also damaging the corneal epithelium. While the mixture of DOL and powder sugar showed an increased corneal thickness and damaged Epithelium. Conclusion: The therapy of corneal edema with osmotically active substances as investigated here is a promising treatment for corneal edema. However, further pharmacological development is needed for application in patients.

Initiating Continuous Renal Replacement Therapy in Patients With Transurethral Resection of Prostate Syndrome: A Case Report

With advances and developments in hysteroscopy, cystoscopy, transurethral resection of bladder tumor, and arthroscopy, transurethral resection of prostate (TURP) syndrome has been increasingly reported. TURP syndrome is often accompanied by severe hyponatremia, fluid overload, and a plasma hypotonic state, resulting in heart failure and pulmonary and cerebral edema. Conventional treatment methods, such as intravenous infusion of hyperosmotic saline, can rapidly reverse the downward trend of serum sodium levels in efforts to prevent and treat cerebral edema. However, this may not be suitable for patients with cardiac and renal insufficiency and may induce central pontine myelinolysis due to the possibility of worsening volume load and difficulty in controlling the correction rate of serum sodium. The patient described in this report presented with severe hyponatremia (sodium<100 mmol/L) combined with intraoperative pulmonary edema; his cardiac function and oxygenation status deteriorated after an intravenous infusion of 3% hypertonic saline. He underwent continuous renal replacement therapy (CRRT) to prevent the progression of multiple–organ edema and cardiac insufficiency. CRRT has demonstrated efficacy in the treatment of chronic hyponatremia in patients with renal failure, and can slowly and continuously correct water-electrolyte imbalance, acid-base imbalance, and volume overload. TURP syndrome with severe hyponatremia and pulmonary edema was diagnosed; accordingly, the patient was treated with 3% hypertonic saline, furosemide, and CRRT, without the development of overt neurological sequelae.

Alleviation of Endoplasmic Reticulum Stress Enhances Human Corneal Epithelial Cell Viability under Hyperosmotic Conditions.

Tear hyperosmolarity plays an essential role in the initiation and progression of dry-eye disease. Under a hyperosmotic environment, corneal epithelial cells experience perturbations in endoplasmic reticulum function that can lead to proinflammatory signaling and apoptosis. In this study, we investigated the effect of tauroursodeoxycholic acid (TUDCA), a chemical chaperone known to protect against endoplasmic reticulum stress, on corneal epithelial cells exposed to hyperosmotic conditions. We found that the expression of the genes involved in the activation of the unfolded protein response and the pro-apoptotic transcription factor DDIT3 were markedly upregulated in patients with Sjögren’s dry-eye disease and in a human model of corneal epithelial differentiation following treatment with hyperosmotic saline. Experiments in vitro demonstrated that TUDCA prevented hyperosmotically induced cell death by reducing nuclear DNA fragmentation and caspase-3 activation. TUDCA supplementation also led to the transcriptional repression of CXCL8 and IL5, two inflammatory mediators associated with dry-eye pathogenesis. These studies highlight the role of hyperosmotic conditions in promoting endoplasmic reticulum stress in the cornea and identify TUDCA as a potential therapeutic agent for the treatment of dry-eye disease.

Effect of extracellular hyperosmolality during normothermia and hyperthermia on the autophagic response in peripheral blood mononuclear cells from young men.

Heat-stress-induced dehydration is associated with extracellular hyperosmolality. To counteract the associated stress, cells employ cytoprotective mechanisms, including autophagy; however, the autophagic response to hyperosmotic stress has yet to be evaluated in humans. Thus, we investigated autophagy and associated cellular stress pathways [the heat shock response (HSR), apoptosis, and the acute inflammatory response] to isosmotic and hyperosmotic conditions with and without hyperthermia in 12 young men (mean [SD]; 25 [5] yr). Participants received a 90-min intravenous infusion of either isosmotic (ISO; 0.9% NaCl; serum osmolality of 293 [4] mosmol/kgH2O) or hyperosmotic (HYP; 3.0% NaCl; 300 [6] mosmol/kgH2O) saline, followed by passive whole body heating using water perfused suit to increase esophageal temperature by ∼0.8°C. Peripheral blood mononuclear cells were harvested at baseline (preinfusion), postinfusion, and after heating, and changes in protein content were analyzed via Western blotting. Post infusion, the LC3-II/I ratio was higher in HYP compared with ISO infusion (P < 0.001), although no other protein changes were observed (all P > 0.050). Following passive heating, autophagy increased in HYP, as demonstrated by an increase in LC3-II from baseline (P = 0.004) and an elevated LC3-II/I ratio compared with ISO (P = 0.035), and a decrease in p62 when compared with the ISO condition (P = 0.019). This was accompanied by an elevation in cleaved caspase-3 following heating in the HYP condition (P < 0.010); however, the HSR and acute inflammatory response did not change under any condition (all P > 0.050). Taken together, our findings indicate that serum hyperosmolality induces autophagy and apoptotic signaling during mild hyperthermia with minimal autophagic activation during normothermia.NEW & NOTEWORTHY We demonstrate that a physiologically relevant increase in serum osmolality causes minimal activation of the autophagic response. However, the combined stressors of serum hyperosmolality and mild hyperthermia causes activation of both autophagy and apoptotic signaling. Thus, changes in osmotic homeostasis appear to influence the cell's cytoprotective ability during periods of heat stress, highlighting the importance of considering osmotic status when examining autophagic responses in vivo.

Ageing attenuates the effect of extracellular hyperosmolality on whole-body heat exchange during exercise-heat stress.

In humans, hypohydration attenuates sweat secretion and attenuates whole-body heat loss, probably to mitigate further fluid losses and thereby support blood pressure regulation. Recently, however, we demonstrated that the hypohydration-mediated reduction in net whole-body heat exchange (evaporative heat loss-dry heat gain) was blunted in middle-aged compared to younger men during moderate exercise in dry heat; albeit, the underpinning mechanisms could not be determined. Here we evaluated the hypothesis that those findings stemmed from a diminished influence of extracellular hyperosmolality on net whole-body heat exchange in middle-aged-to-older compared to young men. Consistent with that hypothesis, extracellular hyperosmolality induced by an intravenous infusion of hypertonic saline (3% NaCl) reduced net heat exchange and augmented rectal temperature to a greater extent in the young compared to middle-aged-to-older men. Thus, age-related differences in the influence of hypohydration on thermoregulatory function appear to be due to blunted sensitivity to hyperosmolality with ageing. We recently demonstrated that sweating-induced hypohydration attenuated whole-body heat dissipation to a greater extent in young compared to middle-aged men during exercise-heat stress. Here, we evaluated whether this divergent response stemmed from an attenuated influence of extracellular hyperosmolality on heat exchange with ageing. To achieve this, ten young (mean(SD): 25(5)years) and ten middle-aged-to-older (61(5)years) men completed two trials involving a 90-min intravenous infusion of isosmotic saline (0.9% NaCl; ISO) or hyperosmotic saline (3.0% NaCl; HYP) followed by 60min of cycling at a fixed metabolic heat production of 250W/m2 (∼50% peak aerobic power) in dry heat (40°C, ∼17% relative humidity). Whole-body net heat exchange (evaporative heat loss-dry heat gain) was measured via direct calorimetry. Rectal temperature was monitored continuously. Heat exchange was attenuated in HYP compared to ISO in the young (233(20) vs. 251(17)W/m2 ; P=0.002) but not older group (229(16) vs. 227(20)W/m2 ; P=0.621). Further, heat exchange was lower in the middle-aged-to-older vs. young men in ISO (P=0.034) but not in HYP (P=0.623). Similarly, end-exercise rectal temperature was greater in HYP relative to ISO in the young (38.3(0.4)°C vs. 37.9(0.3)°C; P=0.015) but not the middle-aged-to-older men (38.3(0.3)°C vs. 38.2(0.2)°C; P=0.652). Compared to the young, rectal temperature was greater in the middle-aged-to-older during ISO (P=0.035) whereas no between-group difference was observed in HYP (P=0.746). Our findings indicate that ageing blunts the effect of extracellular hyperosmolality on thermoregulatory function during exercise-heat stress.

The effect of astaxanthin on inflammation in hyperosmolarity of experimental dry eye model in vitro and in vivo

Hyperosmolarity is pro-inflammatory stress to the ocular surface epithelium associated with dry eye disease (DED). Astaxanthin (AST) is a kind of carotene, which exists in seafood and plays important roles in the amelioration of inflammatory diseases like arteriosclerosis, inflammatory bowel disease, sepsis, rheumatoid arthritis, gastric inflammation, brain inflammatory diseases. The aim of this study was to characterize the protective effect and potential mechanism of AST on DED in vitro and in vivo. Mouse models and human corneal epithelial cell (HCEC) cultures were exposed to hyperosmotic saline solution (HOSS) in in vitro and in vivo experiments, respectively. Experimental subjects were first pretreated with AST, and then the effect of the compound was assessed with clinical evaluation, real-time PCR (RT-PCR), western blot and immunofluorescent staining. We further investigated the possible mechanism of AST in DED by pre-treating with phosphoinositide 3-kinase inhibitor (LY294002). The addition of AST significantly reduced the expression of High-mobility group box 1 (HMGB1), as well as significantly inhibited the increases of TNF-α, IL-1β in a dose-dependent manner, but promoted the expression of phospho-Akt (p-Akt). BALB/c mice in DE group pretreated with AST showed significantly decreased corneal fluorescein staining scores. Moreover, pretreatment with LY294002 could eliminate the effects of AST preconditioning on the decrease of HMGB1. Our study provides evidence that AST could ameliorate DED which may be related to the inhibition of HMGB1, TNF-α, IL-1β, while PI3K/Akt signaling pathway may be involved in the expression of HMGB1 and the protective effect of AST preconditioning.

Autophagy and Heat Shock Protein 70 Expression During Acute Heat Stress in Isosmotic and Hyperosmotic Conditions in Peripheral Blood Mononuclear Cells from Young Adults: Preliminary Data

Heat‐stress induced dehydration is associated with elevations in extracellular osmolality. To counteract the resultant osmotic stress, cells employ several stress response systems including the heat shock protein (HSP) and autophagy pathways. HSPs refold damaged proteins, whereas autophagy acts to degrade damaged cellular components, thereby promoting cellular survival. Accumulating evidence indicates a coordinated response between the HSP and autophagy systems, whereby both systems are activated under conditions of heat stress. However, it remains unknown whether elevated extracellular osmolality augments the HSP and autophagic response to whole‐body heat stress in humans. Thus, the purpose of this study was to examine changes in HSP (HSP70) and autophagy resulting from elevated intravascular osmolality during acute heat exposure. We hypothesized that both autophagy and HSP70 protein expression would be greater under hyperosmotic stress compared to isosmotic conditions. On separate days (separated by ~1 week), six young men (mean [SD]; 27 [5] years) received a 90‐min intravenous infusion with either isosmotic (ISO; 0.9% NaCl; 0.200 mL·kg−1·min−1) or hyperosmotic (HYP; 3.0% NaCl; 0.125 mL·kg−1·min−1) saline, followed by passive whole‐body heating to ~0.8°C above baseline esopha‐‐‐geal temperature using a water perfused suit. Serum osmolality was determined via freeze‐point depression from blood samples at baseline and at the end of passive heating. Peripheral blood mononuclear harvested at these time‐points were analyzed via Western blotting to assess changes in HSP70 protein and proteins indicative of autophagic activity including microtubule associated protein 1 light chain 3 beta‐II (LC3‐II) and p62/sequesterome‐1 (p62). All values were normalized to β‐actin and reported as fold changes relative to baseline. Data are presented as means and standard deviations (SD) and were compared between‐conditions at the end of passive heating using paired (2‐tailed) t‐tests (α=0.05). At the end of heating, serum osmolality was higher during HYP when compared to ISO (298 [SD 3] vs. 291 [SD 3]; p&lt;0.001). During passive heating, LC3‐II was increased (2.20 [SD 2.12] vs. 1.62 [SD 1.47]; p=0.04) and p62 protein was reduced (0.47 [SD 0.33] vs. 1.20 [SD 0.56]; p=0.01) in HYP compared to ISO, indicating greater autophagic activation during HYP. Despite this, HSP70 protein was lower under HYP conditions following heating than ISO (0.72 [SD 0.24] vs. 1.20 [SD 0.19]; p=0.04). Contrary to our hypothesis, these preliminary findings indicate greater reliance on the autophagic response rather than simultaneous activation of HSP70 during acute heat exposure with super‐imposed hyperosmotic stress.Support or Funding InformationNational Science and Engineering Research Council of Canada

Microglia and brain angiotensin type 1 receptors are involved in desensitising baroreflex by intracerebroventricular hypertonic saline in male Sprague-Dawley rats.

High salt diet alters cardiovascular control by increasing concentration of sodium ions (Na+) in cerebrospinal fluid (CSF) and is a risk factor for hypertension. Hypernatremic conditions activate microglia and upregulate renin-angiotensin system in the brain. Thus, we checked if chronic elevation of CSF Na+ affects neural control of circulatory system via microglia and brain angiotensin type 1 receptors (AT1Rs). Normotensive adult male Sprague-Dawley rats received two-week intracerebroventricular (ICV) infusion of either isoosmotic saline (0.9% NaCl); hyperosmotic saline (5% NaCl); 5% NaCl with minocycline - inhibitor of microglia; 5% NaCl with losartan - AT1R blocker. Fluid intake, urine output, and urinary Na+ excretion were measured before and during ICV infusions. At the end of ICV infusions, blood pressure and heart rate were recorded in awake rats at rest, in response to acute air jet stressor, during pharmacological evaluation of baroreflex, and after autonomic ganglia blockade. CSF and blood were collected for evaluation of Na+ concentration. Baroreflex was blunted in rats ICV infused with 5% NaCl. ICV treatment with losartan or minocycline prevented decrease in baroreflex sensitivity. Hemodynamic parameters at rest, in response to acute stressor and autonomic ganglia blockade were similar in all groups. Neither treatment affected water intake, urine output and urinary Na+ excretion. ICV infusion of 5% NaCl resulted in higher concentration of Na+ in CSF than in control group (0.9% NaCl) and in plasma. Our results indicate that chronic ICV infusion of hyperosmotic saline blunts baroreflex in normotensive rats and this desensitization is mediated by microglia and AT1Rs.

УЧАСТИЕ TRPV1 В РЕГУЛЯЦИИ ПРОДУКЦИИ БИКАРБОНАТОВ И КРОВОТОКА В ЖЕЛУДКЕ ПРИ УВЕЛИЧЕНИИ ПОЛОСТНОЙ КИСЛОТНОСТИ, "Российский физиологический журнал им. И.М. Сеченова"

Целью работы было выяснить, как влияет взаимодействие ванилоидных рецепторов переменного потенциала 1-го типа TRPV1 с эндотелиальной (eNOS) и нейрональной (nNOS) изоформами синтазы оксида азота на секрецию бикарбонатов (HCO) и кровоток в теле желудка на фоне аппликации кислоты на слизистую оболочку желудка (СОЖ). У наркотизированных крыс концентрация HCO3 в полостном перфузате желудка рассчитывалась на основе измеренных значений pH/PCO2, а объемный кровоток в стенке органа регистрировался с помощью лазерной допплеровской флоуметрии. Раздражение СОЖ подкисленным гипертоническим раствором (1 M NaCl, pH 2.0) значительно усиливало секрецию HCO3. Деструкция капсаицин-чувствительных первичных афферентов большими дозами капсаицина устраняла эту реакцию. Блокада TRPV1 не влияла на продукцию HCO и увеличение кровотока, вызванные внутрижелудочной аппликацией кислоты. Тем не менее было показано, что TRPV1 активируют eNOS, что приводит к усилению желудочного кровотока и ослаблению транспорта HCO3 на поверхность СОЖ. На активность nNOS, усиливающую продукцию HCO3, TRPV1 влияния не оказывали.

Osmoregulation and muscle water control of the Amazon fish Oscar Astronotus ocellatus (Cichlidae) when facing salinity stress

ABSTRACTSpecimens of Oscar Astronotus ocellatus from a fish farm were abruptly submitted to salt stress of 14 ppt and 20 ppt, for 3 and 8 h to determine their plasma osmolality. Muscle wet body mass change in vitro was analyzed from control freshwater animals. Fish in 14 ppt presented no osmolality distress even after 8 h. In 20 ppt, a slight increase (10%) in plasma osmolality was observed for both times of exposure when compared to control fish. Muscle slices submitted in vitro to hyper-osmotic saline displayed decreased body mass after 75 min, and slices submitted to hypo-osmotic saline displayed increased body mass after 45 min when compared to control (isosmotic saline). These results reinforce A. ocellatus’s euryhalinity. The fish were able to regulate its internal medium and tolerate 14 ppt, but presented an intense osmotic challenge and low muscle hydration control when facing salinities of 20 ppt.

Ovariectomy uncovers purinergic receptor activation of endothelin-dependent natriuresis.

We recently reported that natriuresis produced by renal medullary salt loading is dependent on endothelin (ET)-1 and purinergic (P2) receptors in male rats. Because sex differences in ET-1 and P2 signaling have been reported, we decided to test whether ovarian sex hormones regulate renal medullary ET-1 and P2-dependent natriuresis. The effect of medullary NaCl loading on Na+ excretion was determined in intact and ovariectomized (OVX) female Sprague-Dawley rats with and without ET-1 or P2 receptor antagonism. Isosmotic saline (284 mosmol/kgH2O) was infused in the renal medullary interstitium of anesthetized rats during a baseline urine collection period, followed by isosmotic or hyperosmotic saline (1,800 mosmol/kgH2O) infusion. Medullary NaCl loading significantly enhanced Na+ excretion in intact and OVX female rats. ETA+B or P2 receptor blockade did not attenuate the natriuretic effect of medullary NaCl loading in intact females, whereas ETA+B or P2 receptor blockade attenuated the natriuretic response to NaCl loading in OVX rats. Activation of medullary P2Y2 and P2Y4 receptors by UTP infusion had no significant effect in intact females but enhanced Na+ excretion in OVX rats. Combined ETA+B receptor blockade significantly inhibited the natriuretic response to UTP observed in OVX rats. These data demonstrate that medullary NaCl loading induces ET-1 and P2-independent natriuresis in intact females. In OVX, activation of medullary P2 receptors promotes ET-dependent natriuresis, suggesting that ovarian hormones may regulate the interplay between the renal ET-1 and P2 signaling systems to facilitate Na+ excretion.

Mesodesma mactroides Gill Cells Exposed to Copper: Does Hyposmotic Saline Increase Cytotoxicity or Cellular Defenses?

Gill cells of filter feeding mollusks have cellular defense mechanisms, such as multixenobiotic resistance (MXR), that allow them to extrude possible contaminants. To analyze the cytotoxicity and cellular defenses of gills in the clam Mesodesma mactroides, gill cells were exposed to copper in both iso- and hyposmotic solutions. Analysis of MXR activity by fluorescence microscopy showed that hyposmotic saline activated defenses, whereas the presence of copper in isosmotic solution inhibited the activation of defenses. Cell viability was decreased in cells exposed to copper in isosmotic saline, but not in cells exposed to hyposmotic saline. We conclude that when cells cannot defend themselves due to decreased MXR, cell death occurs. In addition, gill cells under hyposmotic conditions have a greater capacity for defense and a lower rate of cellular mortality than when they are maintained under isosmotic conditions.

Abstract 095: Ovariectomy Restores Purinergic Receptor Activation of Endothelin-dependent Natriuresis

We have recently reported interplay between renal medullary endothelin-1 (ET-1) and purinergic (P2) systems, which play central roles in controlling Na + homeostasis in male rats. Evidence suggests that sex hormones regulate ET-1 and P2 systems. To test whether activation of the renal medullary P2 receptors promotes ET-dependent natriuresis in females and whether ovariectomy (OVX) modulates this potential interaction, we studied the effect of medullary NaCl loading on Na + excretion in adult intact female and OVX SD rats in the presence and absence of P2 or ET receptor antagonism. Isosmotic saline (284 mOsmol/kg H 2 O) was infused into the renal medullary interstitium during a baseline urine collection period, followed by isosmotic or hyperosmotic saline (1800 mOsmol/kg H 2 O) infusion. Blood pressure, renal blood flow, urine Na + , K + and osmolality were measured. Medullary NaCl loading significantly enhanced Na + excretion in intact females and OVX (from 0.8±0.2 to 6.2±1.6 and from 0.7±0.1 to 5.6±0.8 μmol/min, respectively, n=6-8, p&lt;0.05). The natriuretic effect of NaCl loading in intact females was not attenuated by P2 or ET receptor blockade. Whereas, intramedullary infusion of the P2 receptor antagonist, suramin, inhibited the natriuresis induced by medullary NaCl loading in OVX (from 0.4±0.2 to 0.9±0.4 μmol/min, n=6). Additionally, combined ET A/B receptor blockade (ABT-627 + A-192621) abolished the natriuretic response to medullary NaCl load in OVX rats (from 0.2±0.1 to 1.4±0.7 μmol/min, n=4). Activation of medullary purinergic (P2Y 2/4 ) receptors by UTP infusion had no significant effect in intact females, but enhanced Na + excretion in OVX rats (from 0.5±0.1 to 2.3±0.8 μmol/min, n=5, p&lt;0.05). Combined ET A/B receptor blockade significantly inhibited the natriuretic response to UTP observed in OVX rats. These data suggest that increased medullary NaCl loading induces ET-independent and P2-independent natriuresis in intact females. In OVX, activation of medullary P2 receptors promotes ET-dependent natriuresis, similar to our previous findings in male rats and suggests that OVX restores the interplay between the renal ET-1 and purinergic (P2) signaling systems to facilitate Na + excretion. Funded by AHA 15POST25090329 to EYG and P01 HL95499 to DMP

Activation of purinergic receptors (P2) in the renal medulla promotes endothelin-dependent natriuresis in male rats.

Renal endothelin-1 (ET-1) and purinergic signaling systems regulate Na(+) reabsorption in the renal medulla. A link between the renal ET-1 and purinergic systems was demonstrated in vitro, however, the in vivo interaction between these systems has not been defined. To test whether renal medullary activation of purinergic (P2) receptors promotes ET-dependent natriuresis, we determined the effect of increased medullary NaCl loading on Na(+) excretion and inner medullary ET-1 mRNA expression in anesthetized adult male Sprague-Dawley rats in the presence and absence of purinergic receptor antagonism. Isosmotic saline (NaCl; 284 mosmol/kgH2O) was infused into the medullary interstitium (500 μl/h) during a 30-min baseline urine collection period, followed by isosmotic or hyperosmotic saline (1,800 mosmol/kgH2O) for two further 30-min urine collection periods. Na(+) excretion was significantly increased during intramedullary infusion of hyperosmotic saline. Compared with isosmotic saline, hyperosmotic saline infused into the renal medulla caused significant increases in inner medullary ET-1 mRNA expression. Renal intramedullary infusion of the P2 receptor antagonist suramin inhibited the increase in Na(+) excretion and inner medullary ET-1 mRNA expression induced by NaCl loading in the renal medulla. Activation of medullary P2Y2/4 receptors by infusion of UTP increased urinary Na(+) excretion. Combined ETA and ETB receptor blockade abolished the natriuretic response to intramedullary infusion of UTP. These data demonstrate that activation of medullary P2 receptors promotes ET-dependent natriuresis in male rats, suggesting that the renal ET-1 and purinergic signaling systems interact to efficiently facilitate excretion of a NaCl load.

Plasma hyperosmolality attenuates skin sympathetic nerve activity during passive heat stress in humans.

Plasma hyperosmolality delays the onset for sweat production and cutaneous vasodilatation during heat stress in humans; however, the mechanism by which hyperosmolality exerts this effect remains unknown. This study examined if plasma hyperosmolality exerts a central and/or peripheral modulation of thermoregulatory function in humans. The main findings are that plasma hyperosmolality delays the increase in skin sympathetic nerve activity during whole-body passive heat stress in humans. In contrast, local intradermal infusion of hyperosmotic saline did not affect sweating or cutaneous vasodilatation. These results suggest that plasma hyperosmolality delays the onset threshold for sweating and cutaneous vasodilatation by inhibiting efferent thermoregulatory activity in humans. In humans, plasma hyperosmolality delays the onset of sweating and cutaneous vasodilatation during heat stress. However, it remains unknown if hyperosmolality exerts this effect through a central (i.e. CNS) and/or peripheral (i.e. effector organ) modulation of thermoregulatory activity. We examined if intravenous infusion of hyperosmotic saline affects skin sympathetic nerve activity (SSNA) during whole-body passive heating in healthy humans. Furthermore, we examined if local intradermal infusion of hyperosmotic saline affects sweating and cutaneous vasodilatation during passive heating. Following intravenous infusion of either 0.9% (ISO) or 3.0% (HYPER) NaCl saline, 12 subjects were passively heated until core temperature increased by ∼0.6°C. During each condition, sweating and cutaneous vascular conductance were measured over two intradermal microdialysis probes, one perfused with ISO saline and the other with HYPER saline. Intravenous infusion of HYPER saline increased plasma osmolality (294 ± 3 to 316 ± 5 mOsm kg(-1) H2O, P ≤ 0.01), which remained greater than ISO throughout heating. Plasma hyperosmolality delayed the mean body temperature onset of sweating (+1.24 ± 0.18 vs. +1.60 ± 0.18°C, P ≤ 0.01) and cutaneous vasodilatation (+1.15 ± 0.18 vs. +1.53 ± 0.22°C, P ≤ 0.01), and attenuated the increase in SSNA during heating (+147 ± 178 vs. +427 ± 281%, P ≤ 0.01). Intradermal infusion of HYPER saline increased baseline cutaneous vascular conductance (P ≤ 0.01), which did not increase further during the subsequent heating period (P = 0.11). In contrast, intradermal infusion of HYPER saline did not affect sweating (P = 0.99). These results provide direct evidence that plasma hyperosmolality exerts a central modulatory effect governing efferent thermoregulatory activity in humans.

Tolerance of high and low salinity in the intertidal gastropodStramonita brasiliensis(Muricidae): behaviour and maintenance of tissue water content

Stramonita brasiliensis is a marine intertidal gastropod, an active predator, constantly exposed to salinity variations in its intertidal rocky habitat. The goal of this study was to investigate whether the species would handle salinity increase as well as salinity decrease, given the general paucity of data on molluscs in hypersaline seawater. Snails from Santa Catarina State, southern Brazil, were exposed to abrupt (from full-strength seawater to freshwater or to 70 psu) or gradual (full-strength seawater down to freshwater or up to 70 psu) salinity challenges. The isolated muscular foot was challenged with hypoand hyperosmotic shocks, both of 50% with respect to the isosmotic control. Behaviour (release from the substrate and operculum closure), mantle cavity water and haemolymph osmo-ionic concentrations, and tissue hydration were then assayed. When abruptly exposed, snails remained open and active in hypersaline seawater (until 55 psu), but tended to close the operculum upon seawater dilution (salinity 15 psu). In agreement, tissues swelled more in hyposmotic saline than they shrank in hyperosmotic saline. The great euryhalinity displayed by this snail may contribute to its widespread distribution on intertidal rocky coasts.

Sex Differences in Renal Inner Medullary ET‐1 Gene Expression Levels with Increasing Medullary Osmolality

We previously reported that exposing the renal medulla to hyperosmotic solutions enhances urinary ET‐1 excretion in male rats. We have also seen that female rats have a more robust renal ET‐1 natriuretic capacity compared to males. Therefore, we determined whether intramedullary infusion of hyperosmotic saline has a differential effect on ET‐1 gene expression in inner medullas from anesthetized male and female rats. Isosmotic saline (284 mosmol/kgH2O) was infused into the renal medullary interstitium (500 μl/h) during a 60 min equilibration period and 30 min baseline urine collection period, followed by isosmotic or hyperosmotic saline (1800 mosmol/kgH2O) for two further 30 min periods. At the end of each experiment, the renal inner medulla was isolated for ET‐1 mRNA assessment using the real‐time quantitative polymerase chain reaction (RT‐PCR) technique. Compared with isosmotic saline, hyperosmotic saline infused in the renal medulla caused significant increases in urine flow in both male and female rats (from 6.3±0.9 to 12.2±1.7 and from 8.2±1.3 to 15.0±2.8 μl/min, respectively). Sodium excretion was also significantly enhanced, in both males and females (from 0.6±0.1 to 8.3±1.9 and from 0.9±0.2 to 5.8±1.0 μl/min, respectively). Inner medullary ET‐1 expression in male rats was significantly increased after 60 min of medullary interstitial infusion of hyperosmotic saline (from 1.0± 0.1 to 1.4± 0.1 fold change, p&lt;0.05). In contrast, ET‐1 expression did not change with medullary hyperosmotic infusion in females. These data indicate that an increase in renal inner medullary ET‐1 could play a more important role in the diuretic and natriuretic response to intramedullary hyperosmolality in male compared to female rats.

TRPV4 Channels Enable Na,K‐ATPase Activity in the Lens Epithelium to Respond to a Remote Stimulus

Na,K-ATPase activity is low or absent in the fiber cells that make up most of the eye lens. Ion and water homeostasis of the entire structure is made possible by Na,K-ATPase activity in a single epithelial cell layer at the anterior lens surface. Previously we have shown Src family tyrosine kinase-dependent changes of Na,K-ATPase activity in the epithelium. Here we provide evidence that the epithelium is able to increase Na,K-ATPase activity in response to a remote stimulus in the fiber mass. The posterior pole of intact pig lenses was subjected to localized damage either by a 2.5 mm diameter freeze-thaw wound or by injection of 5 µl of hyperosmotic saline just beneath the surface. After 2-30 min the epithelium was removed from the lens, homogenized and the rate of ouabain-sensitive ATP hydrolysis (Na,K-ATPase activity) was determined. Src family tyrosine kinase (SFK) activation (phosphorylation) was assessed by Western blot. Localized damage to lens fibers at posterior pole caused SFK activation and a significant increase in Na,K-ATPase activity in the epithelium that occurred within minutes. Na,K-ATPase activity almost doubled. The increase in Na,K-ATPase activity was prevented by the SFK inhibitor PP2 (10 µM). Importantly, the SFK phosphorylation response as well as the increase in Na,K-ATPase activity were suppressed by a TRPV4 channel antagonist, HC067947 (10 µM). By Western blot, TRPV4 was detected in the epithelium but not fiber cells. The findings point to a TRPV4-dependent mechanism in the anterior epithelium that enables the cells to sense remote damage in the fiber mass and respond by activating SFK and increasing Na,K-ATPase activity.

The use of hyperosmotic saline for chondroprotection: implications for orthopaedic surgery and cartilage repair

Articular cartilage may experience iatrogenic injury during routine orthopaedic/arthroscopic procedures. This could cause chondrocyte death, leading to cartilage degeneration and posttraumatic osteoarthritis. In an in vitro cartilage injury model, chondrocyte death was reduced by increasing the osmolarity of normal saline (NS), the most commonly-used irrigation solution. Here, we studied the effect of hyperosmolar saline (HS) on chondrocyte viability and cartilage repair in an in vivo injury model. Cartilage injury was induced by a single scalpel cut along the patellar groove of 8 week old rats in the absence of irrigation or with either NS (300 mOsm) or HS (600 mOsm). The percentage of cell death (PCD) within the injured area was assessed using confocal microscopy. Repair from injury was evaluated by histology/immunostaining, and inflammatory response by histology, cytokine array analysis and ELISA (enzyme-linked immunosorbent assay). The PCD in saline-irrigated joints was increased compared to non-irrigated (NI) joints [PCD = 20.8% (95%CI; 14.5, 27.1); PCD = 9.14% (95%CI; 6.3, 11.9); P = 0.0017]. However, hyperosmotic saline reduced chondrocyte death compared to NS (PCD = 10.4% (95%CI; 8.5, 12.3) P = 0.0024). Repair score, type II collagen and aggrecan levels, and injury width, were significantly improved with hyperosmotic compared to NS. Mild synovitis and similar changes in serum cytokine profile occurred in all operated joints irrespective of experimental group. Hyperosmotic saline significantly reduced the chondrocyte death associated with scalpel-induced injury and enhanced cartilage repair. This irrigation solution might be useful as a simple chondroprotective strategy and may also reduce unintentional cartilage injury during articular reconstructive surgery and promote integrative cartilage repair, thereby reducing the risk of posttraumatic osteoarthritis.