Angiotensin-(1-7) serves as an aquaretic by increasing water intake and diuresis in association with downregulation of aquaporin-1 during pregnancy in rats

We previously demonstrated that kidney and urine levels of angiotensin-(1-7) [ANG-(1-7)] were increased in pregnancy. To explore the role of ANG-(1-7) on fluid and electrolyte homeostasis during pregnancy, we evaluated the effect of the ANG-(1-7) antagonist D-alanine-[ANG-(1-7)] (A-779) on kidney function. Virgin and pregnant rats received infusion of vehicle or A-779 (48 microg.kg(-1).h(-1)) for 8 days by osmotic minipumps. Metabolic studies were done on treatment day 7-8. Virgin and pregnant rats at day 15 and 19 were killed, and blood and kidneys were collected. Kidneys were prepared for Western blot analysis for aquaporin-1 (AQP1) and aquaporin-2. In virgin female rats, A-779 increased urine volume and decreased urinary osmolality and AQP1 with no change in water intake. In 19-day pregnant rats, A-779 significantly decreased water intake and urine volume and increased urinary osmolality and kidney AQP1 expression. Only in late gestation did A-779 treatment decrease the difference between intake and output (balance). A-779 treatment increased plasma vasopressin in late gestation but did not change vasopressin in virgins. In virgin and pregnant animals, A-779 administration had no effect on blood pressure, plasma volume, blood volume, or urinary electrolytes. These results suggest that ANG-(1-7) produces antidiuresis associated with upregulation of AQP1 in virgin rats, whereas ANG-(1-7) produces diuresis in late gestation with downregulation of AQP1. ANG-(1-7) contributes to the enhanced water intake during pregnancy, allowing maintenance of the normal volume-expanded state despite diuresis produced in part by decreased AVP and AQP1.     

Hepatic glutathione and nitric oxide are critical for hepatic insulin-sensitizing substance action

We tested the hypothesis that hepatic nitric oxide (NO) and glutathione (GSH) are involved in the synthesis of a putative hormone referred to as hepatic insulin-sensitizing substance HISS. Insulin action was assessed in Wistar rats using the rapid insulin sensitivity test (RIST). Blockade of hepatic NO synthesis with N(G)-nitro-l-arginine methyl ester (l-NAME, 1.0 mg/kg intraportal) decreased insulin sensitivity by 45.1 +/- 2.1% compared with control (from 287.3 +/- 18.1 to 155.3 +/- 10.1 mg glucose/kg, P < 0.05). Insulin sensitivity was restored to 321.7 +/- 44.7 mg glucose/kg after administration of an NO donor, intraportal SIN-1 (5 mg/kg), which promotes GSH nitrosation, but not after intraportal sodium nitroprusside (20 nmol x kg(-1) x min(-1)), which does not nitrosate GSH. We depleted hepatic GSH using the GSH synthesis inhibitor l-buthionine-[S,R]-sulfoximine (BSO, 2 mmol/kg body wt ip for 20 days), which reduced insulin sensitivity by 39.1%. Insulin sensitivity after l-NAME was not significantly different between BSO- and sham-treated animals. SIN-1 did not reverse the insulin resistance induced by l-NAME in the BSO-treated group. These results support our hypothesis that NO and GSH are essential for insulin action.     

Effect of nonsteroidal anti-inflammatory drugs with varying extent of COX-2-COX-1 selectivity on urinary sodium and potassium excretion in the rat

Nonsteroidal anti-inflammatory drugs (NSAIDs) have different selectivity to inhibit cyclooxygenase-1 (COX-1) and COX-2. Treatment with NSAIDs has been associated with kidney side effects. We compared the effect of a selected group of NSAIDs with different COX-2--COX-1 selectivities on urinary sodium and potassium excretion in rats. Each treatment with rofecoxib, celecoxib, meloxicam, diclofenac, and flurbiprofen (30, 120, 9, 30, and 125 mg/kg, respectively) and placebo was administered orally once daily for 4 days. Urine was collected 0-8 h after each dose. Urinary sodium and potassium excretion and urine flow rate were compared with placebo. As compared with placebo, rofecoxib, celecoxib, diclofenac, and flurbiprofen significantly reduced excretion rate of sodium (rofecoxib, 0.28 +/- 0.02 vs. 0.41 +/- 0.03; celecoxib, 0.23 +/- 0.03 vs. 0.48 +/- 0.04; diclofenac, 0.09 +/- 0.02 vs. 0.46 +/- 0.03; and flurbiprofen, 0.11 +/- 0.02 vs. 0.47 +/- 0.02 micromol/(min x 100 g)) and potassium (rofecoxib, 0.55 +/- 0.04 vs. 0.68 +/- 0.04; celecoxib, 0.50 +/- 0.06 vs. 0.72 +/- 0.06; diclofenac, 0.26 +/- 0.05 vs. 0.67 +/- 0.04; and flurbiprofen, 0.35 +/- 0.05 vs. 0.62 +/- 0.03 micromol/ (min x 100 g)). Rofecoxib and flurbiprofen significantly reduced urine flow rate. Meloxicam had no significant effect on either sodium and potassium excretion or on the urine flow rate. At the examined dosage level, no relationship was found between reported COX-2--COX-1 selectivity and urinary electrolytes excretion.     

AT1a receptor signaling is required for basal and water deprivation-induced urine concentration in AT1a receptor-deficient mice

It is well recognized that ANG II interacts with arginine vasopressin (AVP) to regulate water reabsorption and urine concentration in the kidney. The present study used ANG II type 1a (AT(1a)) receptor-deficient (Agtr1a(-/-)) mice to test the hypothesis that AT(1a) receptor signaling is required for basal and water deprivation-induced urine concentration in the renal medulla. Eight groups of wild-type (WT) and Agtr1a(-/-) mice were treated with or without 24-h water deprivation and 1-desamino-8-d-AVP (DDAVP; 100 ng/h ip) for 2 wk or with losartan (10 mg/kg ip) during water deprivation. Under basal conditions, Agtr1a(-/-) mice had lower systolic blood pressure (P < 0.01), greater than threefold higher 24-h urine excretion (WT mice: 1.3 ± 0.1 ml vs. Agtr1a(-/-) mice: 5.9 ± 0.7 ml, P < 0.01), and markedly decreased urine osmolality (WT mice: 1,834 ± 86 mosM/kg vs. Agtr1a(-/-) mice: 843 ± 170 mosM/kg, P < 0.01), without significant changes in 24-h urinary Na(+) excretion. These responses in Agtr1a(-/-) mice were associated with lower basal plasma AVP (WT mice: 105 ± 8 pg/ml vs. Agtr1a(-/-) mice: 67 ± 6 pg/ml, P < 0.01) and decreases in total lysate and membrane aquaporin-2 (AQP2; 48.6 ± 7% of WT mice, P < 0.001) and adenylyl cyclase isoform III (55.6 ± 8% of WT mice, P < 0.01) proteins. Although 24-h water deprivation increased plasma AVP to the same levels in both strains, 24-h urine excretion was still higher, whereas urine osmolality remained lower, in Agtr1a(-/-) mice (P < 0.01). Water deprivation increased total lysate AQP2 proteins in the inner medulla but had no effect on adenylyl cyclase III, phosphorylated MAPK ERK1/2, and membrane AQP2 proteins in Agtr1a(-/-) mice. Furthermore, infusion of DDAVP for 2 wk was unable to correct the urine-concentrating defects in Agtr1a(-/-) mice. These results demonstrate that AT(1a) receptor-mediated ANG II signaling is required to maintain tonic AVP release and regulate V(2) receptor-mediated responses to water deprivation in the inner medulla.     

Chronic use of chloroquine disrupts the urine concentration mechanism by lowering cAMP levels in the inner medulla

Chloroquine, a widely used anti-malaria drug, has gained popularity for the treatment of rheumatoid arthritis, systemic lupus erythematosus (SLE), and human immunodeficiency virus (HIV). Unfortunately, chloroquine may also negatively impact renal function for patients whose fluid and electrolyte homeostasis is already compromised by diseases. Chronic administration of chloroquine also results in polyuria, which may be explained by suppression of the antidiuretic response of vasopressin. Several of the transporters responsible for concentrating urine are vasopressin-sensitive including the urea transporters UT-A1 and UT-A3, the water channel aquaporin-2 (AQP2), and the Na(+)-K(+)-2Cl(-) cotransporter (NKCC2). To examine the effect of chloroquine on these transporters, Sprague-Dawley rats received daily subcutaneous injections of 80 mg·kg(-1)·day(-1) of chloroquine for 4 days. Twenty-four hour urine output was twofold higher, and urine osmolality was decreased by twofold in chloroquine-treated rats compared with controls. Urine analysis of treated rats detected the presence chloroquine as well as decreased urine urea and cAMP levels compared with control rats. Western blot analysis showed a downregulation of AQP2 and NKCC2 transporters; however, UT-A1 and UT-A3 abundances were unaffected by chloroquine treatment. Immunohistochemistry showed a marked reduction of UT-A1 and AQP2 in the apical membrane in inner medullary collecting ducts of chloroquine-treated rats. In conclusion, chloroquine-induced polyuria likely occurs as a result of lowered cAMP production. These findings suggest that chronic chloroquine treatment would exacerbate the already compromised fluid homeostasis observed in diseases like chronic kidney disease.     

Altered expression of renal aquaporins and Na(+) transporters in rats treated with L-type calcium blocker

Nifedipine, a calcium antagonist, has diuretic and natriuretic properties. However, the molecular mechanisms by which these effects are produced are poorly understood. We examined kidney abundance of aquaporins (AQP1, AQP2, and AQP3) and major sodium transporters [type 3 Na/H exchanger (NHE-3); type 2 Na-Pi cotransporter (NaPi-2); Na-K-ATPase; type 1 bumetanide-sensitive cotransporter (BSC-1); and thiazide-sensitive Na-Cl cotransporter (TSC)] as well as inner medullary abundance of AQP2, phosphorylated-AQP2 (p-AQP2), AQP3, and calcium-sensing receptor (CaR). Rats treated with nifedipine orally (700 mg/kg) for 19 days had a significant increase in urine output, whereas urinary osmolality and solute-free water reabsorption were markedly reduced. Consistent with this, immunoblotting revealed a significant decrease in the abundance of whole kidney AQP2 (47 +/- 7% of control rats, P < 0.05) and in inner medullary AQP2 (60 +/- 7%) as well as in p-AQP2 abundance (17 +/- 6%) in nifedipine-treated rats. In contrast, whole kidney AQP3 abundance was significantly increased (219 +/- 28%). Of potential importance in modulating AQP2 levels, the abundance of CaR in the inner medulla was significantly increased (295 +/- 25%) in nifedipine-treated rats. Nifedipine treatment was also associated with increased urinary sodium excretion. Consistent with this, semiquantitative immunoblotting revealed significant reductions in the abundance of proximal tubule Na(+) transporters: NHE-3 (3 +/- 1%), NaPi-2 (53 +/- 12%), and Na-K-ATPase (74 +/- 5%). In contrast, the abundance of the distal tubule Na-Cl cotransporter (TSC) was markedly increased (240 +/- 29%), whereas BSC-1 in the thick ascending limb was not altered. In conclusion, 1) increased urine output and reduced urinary concentration in nifedipine-treated-rats may, in part, be due to downregulation of AQP2 and p-AQP2 levels; 2) CaR might be involved in the regulation of water reabsorption in the inner medulla collecting duct; 3) reduced expression of proximal tubule Na(+) transporters (NHE-3, NaPi-2, and Na, K-ATPase) may be involved in the increased urinary sodium excretion; and 4) increase in TSC expression may occur as a compensatory mechanism.     

Renal cortical and medullary blood flow responses to L-NAME and ANG II in wild-type, nNOS null mutant, and eNOS null mutant mice

Experiments in wild-type (WT; C57BL/6J) mice, endothelial nitric oxide synthase null mutant [eNOS(-/-)] mice, and neuronal NOS null mutant [nNOS(-/-)] mice were performed to determine which NOS isoform regulates renal cortical and medullary blood flow under basal conditions and during the infusion of ANG II. Inhibition of NOS with N(omega)-nitro-l-arginine methyl ester (l-NAME; 50 mg/kg iv) in Inactin-anesthetized WT and nNOS(-/-) mice increased arterial blood pressure by 28-31 mmHg and significantly decreased blood flow in the renal cortex (18-24%) and the renal medulla (13-18%). In contrast, blood pressure and renal cortical and medullary blood flow were unaltered after l-NAME administration to eNOS(-/-) mice, indicating that NO derived from eNOS regulates baseline vascular resistance in mice. In subsequent experiments, intravenous ANG II (20 ng x kg(-1) x min(-1)) significantly decreased renal cortical blood flow (by 15-25%) in WT, eNOS(-/-), nNOS(-/-), and WT mice treated with l-NAME. The infusion of ANG II, however, led to a significant increase in medullary blood flow (12-15%) in WT and eNOS(-/-) mice. The increase in medullary blood flow following ANG II infusion was not observed in nNOS(-/-) mice, in WT or eNOS(-/-) mice pretreated with l-NAME, or in WT mice administered the nNOS inhibitor 5-(1-imino-3-butenyl)-l-ornithine (1 mg x kg(-1) x h(-1)). These data demonstrate that NO from eNOS regulates baseline blood flow in the mouse renal cortex and medulla, while NO produced by nNOS mediates an increase in medullary blood flow in response to ANG II.     

Therapeutic effects of PG201, an ethanol extract from herbs, through cartilage protection on collagenase-induced arthritis in rabbits

In order to assess the therapeutic effects of PG201 (an ethanol extract from herbs) on osteoarthritis, we investigated whether PG201 could suppress the disease progression of collagenase-induced arthritis (CNIA) in rabbits. The right knees of rabbits were injected intra-articularly with collagenase, and the rabbits were orally treated with distilled water (DW), PG201 (200 mg/kg) or diclofenac (DCF, 10 mg/kg) once a day for 8 weeks. Oral administration of PG201 significantly suppressed the stiffness and bone space narrowing. Cartilage erosion and GAG release (p<0.01) were considerably reduced in the knee joints. As well, the mRNA expression of matrix degradation enzymes including MMP-1, -3, and -13 was decreased. On the contrary, the concentrations of TIMP-2 in the synovial fluids were considerably amplified in the PG201 treated group (p<0.01), but not in the DCF treated group. The pathologic inflammatory molecules involved in cartilage destruction such as IL-1beta, PGE2, and NO were also diminished by PG201. Taken together, these results indicate that PG201 has therapeutic effects on CNIA through the prominent protection of cartilage. PG201 indeed has great potential as a form of treatment for osteoarthritis.     

Role of 20-hydroxyeicosatetraenoic acid in the renal and vasoconstrictor actions of angiotensin II

The present study examined the effects of ANG II on the renal synthesis of 20-hydroxyeicosatetraenoic acid (20-HETE) and its contribution to the renal vasoconstrictor and the acute and chronic pressor effects of ANG II in rats. ANG II (10(-11) to 10(-7) mol/l) reduced the diameter of renal interlobular arteries treated with inhibitors of nitric oxide synthase and cyclooxygenase, lipoxygenase, and epoxygenase by 81 +/- 8%. Subsequent blockade of the synthesis of 20-HETE with 17-octadecynoic acid (1 micromol/l) increased the ED(50) for ANG II-induced constriction by a factor of 15 and diminished the maximal response by 61%. Graded intravenous infusion of ANG II (5-200 ng/min) dose dependently increased mean arterial pressure (MAP) in thiobutylbarbitol-anesthetized rats by 35 mmHg. Acute blockade of the formation of 20-HETE with dibromododecenyl methylsulfimide (DDMS; 10 mg/kg) attenuated the pressor response to ANG II by 40%. An intravenous infusion of ANG II (50 ng. kg(-1). min(-1)) in rats for 5 days increased the formation of 20-HETE and epoxyeicosatrienoic acids (EETs) in renal cortical microsomes by 60 and 400%, respectively, and increased MAP by 78 mmHg. Chronic blockade of the synthesis of 20-HETE with intravenous infusion of DDMS (1 mg. kg(-1). h(-1)) or EETs and 20-HETE with 1-aminobenzotriazole (ABT; 2.2 mg. kg(-1). h(-1)) attenuated the ANG II-induced rise in MAP by 40%. Control urinary excretion of 20-HETE averaged 350 +/- 23 ng/day and increased to 1,020 +/- 105 ng/day in rats infused with ANG II (50 ng. kg(-1). min(-1)) for 5 days. In contrast, urinary excretion of 20-HETE only rose to 400 +/- 40 and 600 +/- 25 ng/day in rats chronically treated with ANG II and ABT or DDMS respectively. These results suggest that acute and chronic elevations in circulating ANG II levels increase the formation of 20-HETE in the kidney and peripheral vasculature and that 20-HETE contributes to the acute and chronic pressor effects of ANG II.     

Neurophysiological basis for neurogenic-mediated articular cartilage anabolism alteration

This study was designed to investigate the pathways involved in neurogenic-mediated articular cartilage damage triggered by a nonsystemic distant subcutaneous or intra-articular inflammation. The cartilage damage was assessed 24 h after subcutaneous or intra-articular complete Freund's adjuvant (CFA) injection measuring patellar proteoglycan (PG) synthesis (ex vivo [Na(2)(35)SO(4)] incorporation) in 96 Wistar rats. Unilateral subcutaneous or intra-articular injection of CFA induced significant decrease (25-29%) in PG synthesis in both patellae. Chronic administration of capsaicin (50 mg. kg(-1). day(-1) during 4 days), which blunted the normal response of C fiber stimulation, prevented the bilateral significant decrease in cartilage synthesis. Similarly, intrathecal injection of MK-801 (10 nmol/day during 5 days), which blocked the glutamatergic synaptic transmission at the dorsal horn of signal originating in primary afferent C fibers, eliminated the CFA-induced PG synthesis decrease in both patellae. Chemical sympathectomy, induced by guanethidine (12.5 mg. kg(-1). day(-1) during 6 wk), also prevented PG synthesis alteration. Finally, compression of the spinal cord at the T3-T5 level had a similar protective effect on the reduction of [Na(2)(35)SO(4)] incorporation. It is concluded that the signal that triggers articular cartilage synthesis damage induced by a distant local inflammation 1) is transmitted through the afferent C fibers, 2) makes glutamatergic synaptic connections with the preganglionic neurons of the sympathetic system, and 3) involves spinal and supraspinal pathways.     

Influence of nitric oxide synthase inhibitors on the vasopressin-induced pituitary-adrenocortical activity and hypothalamic catecholamine levels.

In the present study the role of endogenous nitric oxide (NO) in the vasopressin-induced ACTH and corticosterone secretion was investigated in conscious rats. Vasopressin (AVP 5 microg/kg i.p.) considerably augmented ACTH and corticosterone secretion. L-arginine (120 and 300 mg/kg i.p.) did not significantly alter the AVP-induced secretion of those hormones. Nitric oxide synthase (NOS) blockers N(omega)-nitro-L-arginine (L-NNA) and its methyl ester (L-NAME) given i.p. 15 min before AVP markedly increased the AVP-induced ACTH secretion. L-NNA (2 mg/kg) more potently and significantly increased the AVP-induced ACTH secretion, whereas L-NAME elicited a weaker and not significant effect. Both those NOS antagonists intensified significantly and to a similar extent the AVP-induced corticosterone secretion. L-arginine (120 mg/kg i.p.) reversed the L-NNA-induced rise in the AVP-stimulated ACTH secretion and substantially diminished the accompanying corticosterone secretion. Neither vasopressin alone nor in combination with L-arginine and L-NAME evoked any significant alterations in the hypothalamic noradrenaline and dopamine levels. L-NNA (2 and 10 mg/kg i.p.) elicited a dose dependent and significant decrease in the hypothalamic noradrenaline level. The hypothalamic dopamine level was not significantly altered by any treatment. These results indicate that in conscious rats endogenous NO has an inhibitory influence on the AVP-induced increase in ACTH and corticosterone secretion. L-NNA is significantly more potent than L-NAME in increasing the AVP-induced ACTH secretion. This may be connected with a considerable increase by L-NNA of hypothalamic noradrenergic system activation which stimulates the pituitary-adrenal axis in addition to specific inhibition of NOS.     

Augmented central nitric oxide production inhibits vasopressin release during hemorrhage in acute alcohol-intoxicated rodents

Acute alcohol intoxication (AAI) attenuates the AVP response to hemorrhage, contributing to impaired hemodynamic counter-regulation. This can be restored by central cholinergic stimulation, implicating disrupted signaling regulating AVP release. AVP is released in response to hemorrhage and hyperosmolality. Studies have demonstrated nitric oxide (NO) to play an inhibitory role on AVP release. AAI has been shown to increase NO content in the paraventricular nucleus. We hypothesized that the attenuated AVP response to hemorrhage during AAI is the result of increased central NO inhibition. In addition, we predicted that the increased NO tone during AAI would impair the AVP response to hyperosmolality. Conscious male Sprague-Dawley rats (300-325 g) received a 15-h intragastric infusion of alcohol (2.5 g/kg + 300 mg·kg(-1)·h(-1)) or dextrose prior to a 60-min fixed-pressure hemorrhage (～40 mmHg) or 5% hypertonic saline infusion (0.05 ml·kg(-1)·min(-1)). AAI attenuated the AVP response to hemorrhage, which was associated with increased paraventricular NO content. In contrast, AAI did not impair the AVP response to hyperosmolality. This was accompanied by decreased paraventricular NO content. To confirm the role of NO in the alcohol-induced inhibition of AVP release during hemorrhage, the nitric oxide synthase inhibitor, nitro-l-arginine methyl ester (l-NAME; 250 μg/5 μl), was administered centrally prior to hemorrhage. l-NAME did not further increase AVP levels during hemorrhage in dextrose-treated animals; however, it restored the AVP response during AAI. These results indicate that AAI impairs the AVP response to hemorrhage, while not affecting the response to hyperosmolality. Furthermore, these data demonstrate that the attenuated AVP response to hemorrhage is the result of augmented central NO inhibition.     

Prostaglandin synthesis by the cochlea of the guinea pig. Influence of aspirin, gentamicin, and acoustic stimulation

This study describes the synthesis of prostaglandins (PGs) by the vascular structures of the inner ear (lateral wall = stria vascularis and spiral ligament) in vitro. The main PGs produced were PGI2, PGF2 alpha and PGE2. PGI2 and PGF2 alpha were also found in the perilymph. A 350 mg/kg ip injection of aspirin decreased PG synthesis by the lateral wall and PG levels in perilymph. This effect was reversed after 3 days. Gentamicin (10(-9) to 10(-5) M) decreased significantly and reversibly PG synthesis in vitro, as did 100 mg/kg ip injection. Acoustic stimulation increased ex vivo PGI2 and PGE2 synthesis without modifying PG levels in perilymph. Results suggest that PGs could be one humoral mediator of the cochlear microcirculation homeostasis, and, possibly, of the circulatory disturbances reported after acoustic stimulation. The decreased PG synthesis after gentamicin treatment could account for the angiotoxic component observed in aminoglycoside ototoxicity.     

NO-dependent blood pressure regulation in RGS2-deficient mice

The regulator of G protein signaling (RGS) 2, a GTPase-activating protein, is activated via the nitric oxide (NO)-cGMP pathway and thereby may influence blood pressure regulation. To test that notion, we measured mean arterial blood pressure (MAP) and heart rate (HR) with telemetry in N(omega)-nitro-l-arginine methyl ester (l-NAME, 5 mg l-NAME/10 ml tap water)-treated RGS2-deficient (RGS2(-/-)) and RGS2-sufficient (RGS2(+/+)) mice and assessed autonomic function. Without l-NAME, RGS2(-/-) mice showed during day and night a similar increase of MAP compared with controls. l-NAME treatment increased MAP in both strains. nNOS is involved in this l-NAME-dependent blood pressure increase, since 7-nitroindazole increased MAP by 8 and 9 mmHg (P < 0.05) in both strains. The l-NAME-induced MAP increase of 14-15 mmHg during night was similar in both strains. However, the l-NAME-induced MAP increase during the day was smaller in RGS2(-/-) than in RGS2(+/+) (11 +/- 1 vs. 17 +/- 2 mmHg; P < 0.05). Urinary norepinephrine and epinephrine excretion was higher in RGS2(-/-) than in RGS2(+/+) mice. The MAP decrease after prazosin was more pronounced in l-NAME-RGS2(-/-). HR variability parameters [root mean square of successive differences (RMSSD), low-frequency (LF) power, and high-frequency (HF) power] and baroreflex sensitivity were increased in RGS2(-/-). Atropine and atropine plus metoprolol markedly reduced RMSSD, LF, and HF. Our data suggest an interaction between RGS2 and the NO-cGMP pathway. The blunted l-NAME response in RGS2(-/-) during the day suggests impaired NO signaling. The MAP increases during the active phase in RGS2(-/-) mice may be related to central sympathetic activation and increased vascular adrenergic responsiveness.     

Hexadecane mineralization and denitrification in two diesel fuel-contaminated soils

The effect of nitrate, ammonium and urea on the mineralization of [(14)C]hexadecane (C(16)H(34)) and on denitrification was evaluated in two soils contaminated with diesel fuel. In soil A, addition of N fertilizers did not stimulate or inhibit background hexadecane mineralization (4.3 mg C(16)H(34) kg(-1) day(-1)). In soil B, only NaNO(3) stimulated hexadecane mineralization (0.91 mg C(16)H(34) kg(-1) day(-1)) compared to soil not supplemented with any nitrogen nutrient (0.17 mg C(16)H(34) kg(-1) day(-1)). Hexadecane mineralization was not stimulated in this soil by NH(4)NO(3) (0.13 mg C(16)H(34) kg(-1) day(-1)), but the addition of NH(4)Cl or urea suppressed hexadecane mineralization (0.015 mg C(16)H(34) kg(-1) day(-1)). Addition of 2 kPa C(2)H(2) did not inhibit the mineralization process in either soil. Denitrification occurred in both soils studied when supplemented with NaNO(3) and NH(4)NO(3), but was not detected with other N sources. Denitrification started after a longer lag in soil A (10 days) than in soil B (4 days). In soil A microcosms supplemented with NaNO(3) or NH(4)NO(3), rates of denitrification were 20.6 and 13.6 mg NO(3)(-) kg(-1) day(-1), respectively, and in soil B, they were 18.5 and 12.5 mg NO(3)(-) kg(-1) day(-1), respectively. We conclude that denitrification may lead to a substantial loss of nitrate, making it unavailable to the mineralizing bacterial population. Nitrous oxide was an important end-product accounting for 30-100% of total denitrification. These results indicate the need for preliminary treatability studies before implementing full-scale treatment processes incorporating commercial fertilizers.     

Sex differences in vasopressin V₂ receptor expression and vasopressin-induced antidiuresis

The renal vasopressin V(2) receptor (V(2)R) plays a critical role in physiological and pathophysiological processes associated with arginine vasopressin (AVP)-induced antidiuresis. Because clinical data suggests that females may be more prone to hyponatremia from AVP-mediated antidiuresis, we investigated whether there are sex differences in the expression and function of the renal V(2)R. In normal Sprague-Dawley rat kidneys, V(2)R mRNA and protein expression was 2.6- and 1.7-fold higher, respectively, in females compared with males. To investigate the potential physiological implications of this sex difference, we studied changes in urine osmolality induced by the AVP V(2)R agonist desmopressin. In response to different doses of desmopressin, there was a graded increase in urine osmolality and decrease in urine volume during a 24-h infusion. Females showed greater mean increases in urine osmolality and greater mean decreases in urine volume at 0.5 and 5.0 ng/h infusion rates. We also studied renal escape from antidiuresis produced by water loading in rats infused with desmopressin (5.0 ng/h). After 5 days of water loading, urine osmolality of both female and male rats escaped to the same degree physiologically, but V(2)R mRNA and protein in female kidneys was reduced to a greater degree (-63% and -73%, respectively) than in males (-32% and -48%, respectively). By the end of the 5-day escape period, renal V(2)R mRNA and protein expression were reduced to the same relative levels in males and females, thereby abolishing the sex differences in V(2)R expression seen in the basal state. Our results demonstrate that female rats express significantly more V(2)R mRNA and protein in kidneys than males, and that this results physiologically in a greater sensitivity to V(2)R agonist administration. The potential pathophysiological implications of these results are that females may be more susceptible to the development of dilutional hyponatremia because of a greater sensitivity to endogenously secreted AVP.     

Role of nitric oxide mechanisms in gastric emptying of, and the blood pressure and glycemic responses to, oral glucose in healthy older subjects

The primary aims of this study were to evaluate the effects of the nitric oxide (NO) synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME) on gastric emptying (GE) of, and the blood pressure (BP), glycemic, insulin, and incretin responses to, oral glucose in older subjects. Eight healthy subjects (4 males and 4 females, aged 70.9 +/- 1.3 yr) were studied on two separate days, in double-blind, randomized order. Subjects received an intravenous infusion of either l-NAME (180 mug.kg(-1).h(-1)) or saline (0.9%) at a rate of 3 ml/min for 150 min. Thirty minutes after the commencement of the infusion (0 min), subjects consumed a 300-ml drink containing 50 g glucose labeled with 20 MBq (99m)Tc-sulfur colloid, while sitting in front of a gamma camera. GE, BP (systolic and diastolic), heart rate (HR), blood glucose, plasma insulin, and incretin hormones, glucose-dependant insulinotropic-polypeptide (GIP), and glucagon-like peptide-1 (GLP-1), were measured. l-NAME had no effect on GE, GIP, and GLP-1. Between -30 and 0 min l-NAME had no effect on BP or HR. After the drink (0-60 min), systolic and diastolic BP fell (P < 0.05) and HR increased (P < 0.01) during saline; these effects were attenuated (P < 0.001) by l-NAME. Blood glucose levels between 90 and 150 min were higher (P < 0.001) and plasma insulin were between 15 and 150 min less (P < 0.001) after l-NAME. The fall in BP, increase in HR, and stimulation of insulin secretion by oral glucose in older subjects were mediated by NO mechanisms by an effect unrelated to GE or changes in incretin hormones.     

Prostaglandins mediate suppression of lymphocyte proliferation and cytokine synthesis in acute Trypanosoma cruzi infection

Suppression of host lymphoproliferative responses to mitogens and Ag is characteristically seen during acute infection with the protozoan parasite Trypanosoma cruzi. We investigated the reciprocal regulation of prostaglandins (PG), TNF-alpha, and nitric oxide (NO) production and their effects on cytokine production and lymphoproliferative responses to parasite Ag and to Con A by spleen cells (SC) from T.-cruzi-infected mice. Large amounts of PGE2, TNF-alpha, and NO were produced during infection. TNF-alpha stimulated PG and NO synthesis, while both mediators inhibited TNF-alpha synthesis. Blocking PG also reduced NO synthesis indicating that PG stimulate NO production. Treatment with indomethacin or NMLA stimulated lymphoproliferation on days 6 and 22 of infection; on day 14, when suppression of proliferation and NO production was maximal, combined inhibition of NO and PG production restored parasite Ag specific and Con A proliferative responses. Blocking PG or NO production increased IL-2, IFN-gamma, and TNF-alpha, but not IL-12 production by SC; IL-10 levels were not reduced. Indomethacin-treated infected mice had higher mortality compared to untreated infected animals. The data indicate that PG, together with NO and TNF-alpha, participate in a complex circuit that controls lymphoproliferative and cytokine responses in T. cruzi infection.