Androgen metabolism by hepatic and renal tissues of the fetal rhesus monkey

Liver and kidney from fetal monkeys (day 125 of gestation) were fractionated into low speed pellets, microsomal and cytosolic fractions. Liver cytosols converted as much testosterone (T) to 5 beta-androstane-3 alpha,17 beta-diol (5 beta-diol) at 0 degrees C as at 4 degrees-45 degrees C without exogenous cofactors. The principal product formed from 5 alpha-dihydrotestosterone (5 alpha-DHT) was 5 alpha-diol. A 1000-fold molar excess of radioinert 5 beta- or 5 alpha-DHT inhibited 5 beta-diol formation from [3H]T by cytosols and increased 5 beta-DHT formation. Similarly, using 5 alpha-DHT as substrate, 5 alpha-diol formation was inhibited. Microsomal and low speed pellets with added cofactors formed products which recrystallized with either etiocholanolone or androsterone from [3H]T or [3H]DHT, respectively. Little product was formed without cofactor. Whole liver homogenates produced 5 beta-reduced products from [3H]T in the presence of an NADPH generating system whereas kidney homogenates produced 5 alpha-reduced products. These data provide new information on the capacity of fetal monkey liver and kidney to metabolize androgens. The 3 alpha-reductases are cytosolic. The 5 alpha- and 5 beta-reductases are mostly in the low speed pellet but are sufficiently represented in cytosols to mediate diol formation. The 17-hydroxysteroid dehydrogenases are in the microsomal fraction. Our results suggest that 5 alpha-DHT is the active androgen in fetal liver since testosterone is metabolized to 5 beta-DHT and 5 beta-diol which are inactive androgens.     

Organization and activation of behavior in quail: role of testosterone metabolism

In quail, the hypothalamus enzymatically transforms testosterone (T) into estradiol (E2), 5 alpha-dihydrotestosterone (5 alpha-DHT), and 5 beta-dihydrotestosterone (5 beta-DHT). During the embryonic life, the 5 beta-reductase activity is very high, which probably protects the brain of males from being behaviorally demasculinized by their endogenous T. 5 beta androstanes are inactive androgens. The decrease of 5 beta reductase with age during sexual maturation corresponds to a potentiation of the effects of T as shown by experiments that compared the effects of T and 5 alpha-DHT in adult and young quail. T metabolism is also involved in the activation of male behavior in the adult. T aromatization is probably essential for behavioral activation, but nonaromatizable androgens such as methyltrienolone, and to some extent 5 alpha-DHT, can also stimulate sexual behavior in castrates. These enzymatic activities show a clear neuroanatomical localization and are sexually dimorphic. Males produce more active metabolites (E2, 5 alpha-DHT) than females, which could explain the male's greater sensitivity to T treatments. It thus appears that T metabolism is involved in the differentiation and activation of behavior in quail.     

Frequent occurrence of polyovular follicles in ovaries of mice exposed neonatally to diethylstilbestrol

The occurrence of polyovular follicles (PF) was examined at 10-34 days of age in the ovaries of BALB/cCrgl female mice given five daily injections of 0.1 microgram diethylstilbestrol (DES), 2 micrograms DES, 100 micrograms progesterone (P), 137 micrograms 17 alpha-hydroxyprogesterone caproate (HPC), 20 micrograms testosterone (T), 20 micrograms 5 alpha-dihydrotestosterone (5 alpha-DHT), or oil vehicle alone starting on the day of birth, and of C57BL/Tw females given five neonatal injections of 1 microgram DES, 20 micrograms 17 beta-estradiol (E2), 50 micrograms 5 alpha-DHT, 50 micrograms 5 beta-DHT, or the vehicle alone. Ovaries of 30-day-old C57BL mice given five daily injections of 1 microgram DES starting at 3-25 days of age were also examined. PF incidence (% of PF per ovary) and PF frequency (% of mice with PF) were significantly greater in BALB/c mice receiving injections of DES, P, HPC, and T than in the controls. In DES-treated mice at 34 days, PF incidence (2-13 oocytes/follicle) was 120-340 times higher than in the controls. BALB/c mice treated with T, P, and HPC showed PF incidence (two to four oocytes/follicle) three- to six-fold higher than in the controls. In 30-day-old C57BL mice treated with T, E2, and DES, PF incidence also increased by two- to 50-fold. 5 alpha-DHT and 5 beta-DHT failed to increase PF incidence. PF incidence was significantly increased only when neonatal DES treatment was begun on days 0 to 3, but was reduced when started at days 10-25.(ABSTRACT TRUNCATED AT 250 WORDS)     

Analysis of androgen action on pituitary gonadotropin and prolactin secretion in ewes

To study the role of androgens in the control of gonadotropin and prolactin secretion in ther ewe, we have characterized androgen receptors in pituitary cytosol, and investigated the effect of androgens on pituitary hormone release in vivo and in vitro. High affinity, low capacity receptors, with an affinity for methyltrienolone (R1881) greater than 5 alpha-dihydrotestosterone (5 alpha-DHT) greater than testosterone (T) much greater than androstenedione (A4), estradiol-17 beta (E2) and progesterone (P), were identified in pituitary cytosol. Addition of 1 nM 5 alpha-DHT, but not A4, inhibited luteinizing hormone (LH) release from pituitary cells in vitro, induced by 10(10) to 10(-7) M luteinizing hormone releasing hormone (LHRH). The release of follicle-stimulating hormone (FSH) with 10(-9) M LHRH was inhibited when cells were incubated with 1 nM 5 alpha-DHT. 5 alpha-DHT had no effect when higher or lower doses of LHRH were used. In ovariectomized ewes, neither an i.v. injection of 1 mg, nor intracarotid injections of up to 1 mg, 5 alpha-DHT affected plasma LH, FSH or prolactin levels, despite dose-related increases in plasma 5 alpha-DHT levels. Daily or twice daily i.m. injections of 5 mg 5 alpha-DHT in oil did not affect LH or FSH levels, but daily injections of 20 mg significantly reduced plasma LH levels within 4 days and plasma FSH levels within 6 days. Thus, despite the presence of androgen receptors in the ewe pituitary, we conclude that androgens per se are of minimal importance in the regulation of pituitary LH, FSH and prolactin secretion in the ewe. The low binding affinity of A4 and the lack of its effect on hormone secretion in vitro suggests that A4 may act as an estrogen precursor rather than an androgenic hormone. The function of the pituitary androgen receptor remains to be established.     

Oxytocinergic regulation of cardiovascular function: studies in oxytocin-deficient mice

Oxytocin (OT) has been implicated in the cardiovascular responses to exercise, stress, and baroreflex adjustments. Studies were conducted to determine the effect of genetic manipulation of the OT gene on blood pressure (BP), heart rate (HR), and autonomic/baroreflex function. OT knockout (OTKO -/-) and control +/+ mice were prepared with chronic arterial catheters. OTKO -/- mice exhibited a mild hypotension (102 +/- 3 vs. 110 +/- 3 mmHg). Sympathetic and vagal tone were tested using beta(1)-adrenergic and cholinergic blockade (atenolol and atropine). Magnitude of sympathetic and vagal tone to the heart and periphery was not significantly different between groups. However, there was an upward shift of sympathetic tone to higher HR values in OTKO -/- mice. This displacement combined with unchanged basal HR led to larger responses to cholinergic blockade (+77 +/- 25 vs. +5 +/- 15 beats/min, OTKO -/- vs. control +/+ group). There was also an increase in baroreflex gain (-13.1 +/- 2.5 vs. -4.1 +/- 1.2 beats x min(-1) x mmHg(-1), OTKO -/- vs. control +/+ group) over a smaller BP range. Results show that OTKO -/- mice are characterized by 1) hypotension, suggesting that OT is involved in tonic BP maintenance; 2) enhanced baroreflex gain over a small BP range, suggesting that OT extends the functional range of arterial baroreceptor reflex; and 3) shift in autonomic balance, indicating that OT reduces the sympathetic reserve.     

Induction of vaginal mucification in rats with testosterone and 17beta-hydroxy-5alpha-androstan-3-one.

Based on histological criteria, Kingsley and Bogdanove (3) reported that the benzoate ester of 17beta-hydroxy-5alpha-androstan-3-one (5alpha-DHT), unlike testosterone propionate, is unable to induce vaginal mucification when given subcutaneously to rats. In contrats, Kennedy (4) found in estrogen-pretreated rats that both 5alpha-DHT and testosterone induced vaginal mucification as indicated by increased vaginal sialic acid concentration.To determine if esterification of these androgens altered their ability to induce vaginal mucification, ovariectomized rats, pretreated for 3 days with 0.25 mug estradiol-17beta, were treated for 8 days with either sesame oil or 7 mumoles of testosterone, 5alpha-DHT and their respective propionate and benzoate esters. All treatments except 5-alpha-DHT benzoate increased vaginal weight and vaginal mucification, as assessed histochemically and biochemically. 5alpha-DHT propionate was less effective than 5alpha-DHT while testosterone benzoate, but not propionate was less effective than testosterone. To determine if estrogens are necessary for the vaginal effects of androgens, ovariectomized and ovariectomized-adrenalectomized rats were treated with testosterone or 5alpha-DHT. Adrenalectomy did not significantly affect the vaginal response to either androgen. It is therefore concluded that androgens are capable of inducing vaginal mucification in the absence of estrogens.     

Testosterone metabolism and sexual behavior in the chick

A series of experiments was performed to study the behavioral and physiological activity of testosterone (T) metabolites which are produced by neural tissues of male chicks, i.e., mainly 5α- and 5β-dihydrotestosterone (5α-, 5β-DHT), 5α- and 5β-androstan-3α, 17β-diol (5α-,5β-diol), and 4-androstene-3,17-dione (Δ4). It was found that 5β-reduced androgens alone or in combination with estradiol stimulate juvenile copulation in the chick while they have no detectable effect on all somatic variables which are recorded (testicular weight, plasma LH) with the exception of comb size. On the contrary, comb size was increased by T, Δ4, 5α-DHT, and 5α-diol while testis growth was prevented by T and Δ4 only. There is a good correlation between the anatomical localization of the enzymatic activities which metabolize T and the hormonal dependence of the biological responses: The comb converts T into 5α-reduced compounds which affect comb growth. 5β-Reduction is high in the hypothalamus, a fact which can be related to the sensitivity of sexual behavior to 5β-reduced androgens. This suggests that T metabolism is a very important step in the expression of this hormone's physiological effects.     

Urotensin II-induced hypotensive responses in Wistar-Kyoto (Wky) and spontaneously hypertensive (Shr) rats

Human urotensin II (hU-II) is a potent vasoactive peptide which modulates some of the functions of the cardiovascular and other systems. The in vivo mechanism of action by which hU-II may influence blood pressure in developmental and pathological conditions, is poorly understood. Herein, the blood pressure effects of hU-II (0.1-10 nmol/kg) injected intravenously (i.v.) were studied on ketamine/xylazine anesthetized male WKY and SHR rats aged 4 and 8 weeks. hU-II elicited dose-dependent decreases in mean arterial pressure in both strains of animals. The hypotensive responses to hU-II were, however, significantly higher in SHR rats, independently of age. Four-week-old SHR rats (which are normotensive) were, however, less responsive than their hypertensive 8-week-old counterparts. A series of pharmacological inhibitors were used to identify putative endogenous (endothelial) factors that might account for the hU-II-mediated hypotension in 8-week-old SHR. These include the non-selective nitric oxide synthase inhibitor L-NAME (5 micromol/kg), the non-selective cyclooxygenase inhibitor meclofenamate (16 micromol/kg), the voltage-sensitive and ATP-sensitive K+-channel inhibitors, 4-aminopyridine (5 micromol/kg) and glybenclamide (10 micromol/kg), the cytochrome P450 CYP2C9 inhibitor sulfaphenazole (15 micromol/kg), the cytoskeletal fixation agent phalloidin (15 micromol/kg), the endothelin ETB receptor antagonist BQ-788(35 micromol/kg), the bradykinin B2 receptor antagonist HOE 140 (0.5 micromol/kg), the angiotensin AT2 antagonist PD 123319(10 micromol/kg) and the UT receptor antagonist urantide (10 micromol/kg). These agents were administered i.v. either at 2.5, 10 or 40 min prior hU-II injection (10 nmol/kg). Among these inhibitors, sulfaphenazole and phalloidin were able to reduce hU-II-induced hypotension. This suggests that the vasodepressor effect of hU-II is mediated by UT receptors and relies in part on the release of epoxide related products; increased microvascular permeability may also contribute to the blood pressure lowering effect of hU-II. Since urantide blocks the constrictor effects of hU-II on isolated aorta, but is inactive against the hypotensive action of hU-II in vivo, the results presented in this paper provide, for the first time, evidence for the existence of two different functional sites for hU-II.     

Prolactin directly enhanced Na+/K+- and Ca2+-ATPase activities in the duodenum of female rats

Prolactin has recently been shown to directly stimulate 2 components of the active duodenal calcium transport in female rats, i.e., solvent drag-induced and transcellular-active calcium transport. Since the basolateral Na(+)/K(+)- and Ca(2+)-ATPases, respectively, play important roles in these 2 transport mechanisms, the present study aimed to examine the direct actions of prolactin on the activities of both transporters in sexually mature female Wistar rats. The results showed that 200, 400, and 800 ng/mL prolactin produced a significant increase in the total ATPase activity of duodenal crude homogenate in a dose-dependent manner within 60 min (i.e., from a control value of 1.53 +/- 0.13 to 2.29 +/- 0.21 (p < 0.05), 2.68 +/- 0.19 (p < 0.01), and 3.92 +/- 0.33 (p < 0.001) micromol Pi x (mg protein)(-1) x min(-1), respectively). Activity of Na+/K+-ATPase was increased by 800 ng/mL prolactin from 0.17 +/- 0.03 to 1.18 +/- 0.29 micromol Pi x (mg protein)(-1) x min(-1) (p < 0.01). Prolactin at doses of 400 and 600 ng/mL also significantly increased the activities of Ca(2+)-ATPase in crude homogenate from a control value of 0.84 +/- 0.03 to 1.75 +/- 0.29 (p < 0.05), and 2.30 +/- 0.37 (p < 0.001) micromol Pi x (mg protein)(-1) x min(-1). When the crude homogenate was purified for the basolateral membrane, the Na(+)/K(+)-ATPase activities were elevated 10-fold. In the purified homogenate, 800 ng/mL prolactin increased Na(+)/K(+)-ATPase activity from 1.79 +/- 0.38 to 2.63 +/- 0.44 micromol Pi x (mg protein)(-1) x min(-1) (p < 0.05), and Ca(2+)-ATPase activity from 0.08 +/- 0.14 to 2.03 +/- 0.23 micromol Pi x (mg protein)(-1) x min-1 (p < 0.001). Because the apical calcium entry was the first important step for the transcellular active calcium transport, the brush border calcium uptake was also investigated in this study. We found that, 8 min after being directly exposed to 800 ng/mL prolactin, the brush border calcium uptake into the duodenal epithelial cells was increased from 0.31 +/- 0.02 to 0.80 +/- 0.28 nmol x (mg protein)(-1) (p < 0.05). It was concluded that prolactin directly and rapidly enhanced the brush border calcium uptake as well as the activities of the basolateral Na(+)/K(+)- and Ca(2+)-ATPases in the duodenal epithelium of female rats. These findings explained the mechanisms by which prolactin stimulated duodenal active calcium absorption.     

The role of P(i) in glycolytic inhibition of calcium ion uptake by ELD ascites tumor cells

In contrast to previous investigations at 25 degrees C, glucose was shown to support 45Ca2+ uptake at 37 degrees C in intact ELD ascites tumor cells. Intact ascites tumor cells in vitro accumulated up to 5.0 micromol of 45Ca2+ per g cells dry wt. within 20 min. In the presence of 10.0 mM glucose, intracellular P(i) levels fell from 40.0 micromol x g(-1) cells dry wt. to 20.0 micromol x g(-1) cells dry wt. in 5 min. Intracellular P(i) levels were maintained by 20.0 mM extracellular Tris-P(i). 45Ca2+ uptake was inhibited in P(i)-depleted cells, even though the metabolic rate (as measured by Q(lactate)) and energy state (as measured by ATP levels) were at acceptable levels. Evidence has been presented suggesting that previous reports of glucose inhibition of calcium uptake can be attributed to a competition for available intracellular P(i) between glycolytic processes and the mitochondrial calcium uptake mechanism.     

Cerebrovascular effects of intravenous dopamine infusions in fetal sheep.

Preterm infants are often treated with intravenous dopamine to increase mean arterial blood pressure (MAP). However, there are few data regarding cerebrovascular responses of developing animals to dopamine infusions. We studied eight near-term and eight preterm chronically catheterized unanesthetized fetal sheep. We measured cerebral blood flow and calculated cerebral vascular resistance (CVR) at baseline and during dopamine infusion at 2.5, 7.5, 25, and 75 microg x kg(-1) x min(-1). In preterm fetuses, MAP increased only at 75 microg x kg(-1) x min(-1) (25 +/- 5%), whereas in near-term fetuses MAP increased at 25 microg x kg(-1) x min(-1) (28 +/- 4%) and further at 75 microg x kg(-1) x min(-1) (51 +/- 3%). Dopamine infusion was associated with cerebral vasoconstriction in both groups. At 25 microg x kg(-1) x min(-1), CVR increased 77 +/- 51% in preterm fetuses and 41 +/- 11% in near-term fetuses, and at 75 microg x kg(-1) x min(-1), CVR increased 80 +/- 33% in preterm fetuses and 83 +/- 21% in near-term fetuses. We tested these responses to dopamine in 11 additional near-term fetuses under alpha-adrenergic blockade (phenoxybenzamine, n = 5) and under dopaminergic D(1)-receptor blockade (SCH-23390, n = 6). Phenoxybenzamine completely blocked dopamine's pressor and cerebral vasoconstrictive effects, while D(1)-receptor blockade had no effect. Therefore, in unanesthetized developing fetuses, dopamine infusion is associated with cerebral vasoconstriction, which is likely an autoregulatory, alpha-adrenergic response to an increase in blood pressure.     

5alpha-androstane-3alpha,17beta-diol supports human prostate cancer cell survival and proliferation through androgen receptor-independent signaling pathways: implication of androgen-independent prostate cancer progression

Androgen and androgen receptor (AR) are involved in growth of normal prostate and development of prostatic diseases including prostate cancer. Androgen deprivation therapy is used for treating advanced prostate cancer. This therapeutic approach focuses on suppressing the accumulation of potent androgens, testosterone and 5alpha-dihydrotestosterone (5alpha-DHT), or inactivating the AR. Unfortunately, the majority of patients with prostate cancer eventually advance to androgen-independent states and no longer respond to the therapy. In addition to the potent androgens, 5alpha-androstane-3alpha,17beta-diol (3alpha-diol), reduced from 5alpha-DHT through 3alpha-hydroxysteroid dehydrogenases (3alpha-HSDs), activated signaling may represent a novel pathway responsible for the progression to androgen-independent prostate cancer. Androgen sensitive human prostate cancer LNCaP cells were used to compare 5alpha-DHT and 3alpha-diol activated androgenic effects. In contrast to 5alpha-DHT, 3alpha-diol regulated unique patterns of beta-catenin and Akt expression as well as Akt phosphorylation in parental and in AR-silenced LNCaP cells. More significantly, 3alpha-diol, but not 5alpha-DHT, supported AR-silenced LNCaP cells and AR negative prostate cancer PC-3 cell proliferation. 3alpha-diol-activated androgenic effects in prostate cells cannot be attributed to the accumulation of 5alpha-DHT, since 5alpha-DHT formation was not detected following 3alpha-diol administration. Potential accumulation of 3alpha-diol, as a result of elevated 3alpha-HSD expression in cancerous prostate, may continue to support prostate cancer growth in the presence of androgen deprivation. Future therapeutic strategies for treating advanced prostate cancer might need to target reductive 3alpha-HSD to block intraprostatic 3alpha-diol accumulation.     

Ischemic preconditioning in rats: role of mitochondrial K(ATP) channel in preservation of mitochondrial function

We examined the role of the sarcolemmal and mitochondrial K(ATP) channels in a rat model of ischemic preconditioning (IPC). Infarct size was expressed as a percentage of the area at risk (IS/AAR). IPC significantly reduced infarct size (7 +/- 1%) versus control (56 +/- 1%). The sarcolemmal K(ATP) channel-selective antagonist HMR-1098 administered before IPC did not significantly attenuate cardioprotection. However, pretreatment with the mitochondrial K(ATP) channel-selective antagonist 5-hydroxydecanoic acid (5-HD) 5 min before IPC partially abolished cardioprotection (40 +/- 1%). Diazoxide (10 mg/kg iv) also reduced IS/AAR (36.2 +/- 4.8%), but this effect was abolished by 5-HD. As an index of mitochondrial bioenergetic function, the rate of ATP synthesis in the AAR was examined. Untreated animals synthesized ATP at 2.12 +/- 0.30 micromol x min(-1) x mg mitochondrial protein(-1). Rats subjected to ischemia-reperfusion synthesized ATP at 0.67 +/- 0.06 micromol x min(-1) x mg mitochondrial protein(-1). IPC significantly increased ATP synthesis to 1.86 +/- 0.23 micromol x min(-1) x mg mitochondrial protein(-1). However, when 5-HD was administered before IPC, the preservation of ATP synthesis was attenuated (1.18 +/- 0.15 micromol x min(-1) x mg mitochondrial protein(-1)). These data are consistent with the notion that inhibition of mitochondrial K(ATP) channels attenuates IPC by reducing IPC-induced protection of mitochondrial function.     

Rapid androgen actions on calcium signaling in rat sertoli cells and two human prostatic cell lines: similar biphasic responses between 1 picomolar and 100 nanomolar concentrations

Androgen-induced calcium fluxes and gap junctional intercellular communication (GJIC) were studied in three different cell types. A transient (2-3 min duration) increase in intracellular calcium levels was observed within 20-30 sec of androgen addition, which was followed by a plateau phase with steroid concentrations higher than 1 nM. The kinetics of the calcium responses were similar in immature rat Sertoli cells, which contain normal nuclear receptors; the human prostatic tumor cell line, LNCaP, which contains a mutated nuclear receptor; and the human prostatic cell line, PC3, which does not contain a nuclear receptor. The human A431 tumor cell line did not respond to androgens. Concentrations of testosterone and the synthetic androgen, R1881, between 1-1000 pM induced transient calcium increases with ED(50) values near 1 pM and 1 nM, whereas dihydrotestosterone (DHT) was not active at these concentrations. At concentrations higher than 1 nM, testosterone, R1881, and DHT were equipotent in stimulating an increase in calcium that lasted for more than 10 min, with ED(50) values between 5 and 20 nM. Testosterone covalently bound to albumin was also active, whereas 11 related androstane compounds as well as progesterone and estradiol-17beta were inactive at 1000 nM. The calcium response induced by the three androgens (10 nM) was abolished in all cell types by hydroxyflutamide (1000 nM) and finasteride (1000 nM), but not by cyproterone acetate (1000 nM). The calcium response was also abolished in the absence of extracellular calcium and strongly inhibited by the presence of verapamil. Exposure of the responsive cells to brief (150-sec) pulses of androgens generated calcium responses that were similar to those after continuous exposure. After exposure of Sertoli cells for only 30 sec to 100 nM testosterone, the calcium response lasted for at least 50 min. Although nuclear binding of androgens could be demonstrated, there was no evidence for tight binding to the plasma membrane under similar conditions. When protein synthesis was inhibited, an enhancement of GJIC between rat Sertoli cells, but not between LNCaP cells or PC3 cells, was observed within 15 min of the addition of 10 nM testosterone. Because nuclear androgens are not present in PC3 cells and many functional properties of the responsive system are different from the nuclear receptor in all three cell types, we postulate the existence of an alternative cell surface receptor system with biphasic response characteristics (high and low affinity). The calcium signals are probably coupled to the regulation of gap junctional efficiency between Sertoli cells. The low-affinity receptors may convey complementary androgen signals at elevated local levels such as in the testis, when nuclear receptors are (over)saturated.     

Volume expansion during acute angiotensin II receptor (AT(1)) blockade and NOS inhibition in conscious dogs

The responses to AT(1)-receptor blockade (candesartan 1 mg/kg) and to concomitant volume expansion (saline 35 ml/kg for 90 min) with and without nitric oxide synthase (NOS) inhibition (N(G)-nitro-L-arginine methyl ester 30 microg small middle dot kg(-1) small middle dot min(-1)) were investigated in separate experiments in normal dogs. AT(1) blockade decreased arterial pressure (106 +/- 4 to 96 +/- 5 mmHg) and increased glomerular filtration rate (GFR) by 17% and sodium excretion threefold. NOS inhibition increased arterial pressure (103 +/- 3 to 116 +/- 3 mmHg) and decreased GFR by 21% and reduced sodium excretion by some 80%. Volume expansion increased arterial pressure significantly in all series involving this procedure, most pronounced during combined AT(1) blockade and NOS inhibition (21 +/- 4 mmHg). Volume expansion during AT(1) blockade elicited marked natriuresis (26 +/- 11 to 274 +/- 55 micromol/min) that was severely reduced by concomitant NOS inhibition (10 +/- 3 to 45 +/- 11 micromol/min), but still much larger than that seen with volume expansion during NOS inhibition alone (2 +/- 1 to 23 +/- 7 micromol/min). Volume expansion during AT(1) blockade increased GFR (+30%), less so during combined AT(1) blockade and NOS inhibition (+13%), but it did not increase GFR significantly (P = 0.07) during NOS inhibition alone. Plasma ANG II increased greater than sevenfold with AT(1) blockade and doubled with NOS inhibition (paired t-test, P < 0.05), whereas it decreased by 50-80% during volume expansion irrespective of pretreatment, i.e., during NOS inhibition, volume expansion did not generate subnormal plasma ANG II concentrations. In conclusion, 1) acute AT(1) blockade leads to hyperfiltration, natriuresis, and hyperresponsiveness to volume expansion, 2) these responses are >85% inhibitable by unspecific NOS inhibition, and 3) NOS inhibition alone is followed by increases in plasma ANG II, hypofiltration, and severe antinatriuresis that may be counterbalanced but not overwhelmed by volume expansion. Thus NOS inhibition virtually abolishes the volume expansion natriuresis, at least in part, due to the lack of appropriate inhibition of the renin-angiotensin-aldosterone system.     

Cardiovascular effects of peptide kinin B2 receptor antagonists in rats

Bradykinin (BK) is a vasoactive peptide reputed to play an important role in cardiovascular homeostasis. In this study, we describe the cardiovascular changes (mean blood pressure (BP) and heart rate (HR)) induced by the i.v. administration (left jugular vein) of two selective kinin B2 receptor antagonist, namely icatibant (0.1-1 micromol/kg as a bolus) and MEN1 1270 (0.1-1 micromol/kg as a bolus or 1 micromol/kg infused in 15 or 60 min), in urethane-anaesthetized or conscious rats with an indwelling catheter implanted in the right carotid artery for BP measurements. In conscious rats, icatibant at 0.1 or 0.3 micromol/kg did not change BP but at 0.1 micromol/kg increased HR at 30 min from administration. MEN1 1270 at 0.1 or 0.3 micromol/kg induced a dose-related increase in BP and a concomitant bradycardia (significant at 0.3 micromol/kg) lasting for 5 or 30 min, respectively. Icatibant at 1 micromol/kg induced a slight (P < 0.05) increase in BP that resolved in 5 min and a biphasic tachycardia (peaks at 30 and 90 min from administration). MEN1 1270 at 1 micromol/kg induced a triphasic change in HR (tachycardia in the first 5 min, bradycardia at 30 min, and tachycardia at 90 and 120 min) and a biphasic change in BP (hypotension at 15 min and hypertension at 30 min). The i.v. infusion of MEN1 1270 (1 micromol/kg in 15 or 60 min) produced hypertension, whereas HR was increased only following the 15-min infusion. In urethane-anaesthetized rats, both icatibant and MEN1 1270 (0.1 micromol/kg as a bolus) increased BP and the onset for this effect was correlated with the time course of the antagonism of BK-induced hypotension, where the effect of MEN1 1270 was more rapid than that of icatibant. These results indicate that kinin B2 receptor antagonists can induce acute cardiovascular effects, and the reason for the different haemodynamic profile between icatibant and MEN1 1270 could be putatively attributed to kinetic characteristics.     

Metabolic response to carbohydrate ingestion during exercise in males and females

The present study investigated potential sex-related differences in the metabolic response to carbohydrate (CHO) ingestion during exercise. Moderately endurance-trained men and women (n = 8 for each sex) performed 2 h of cycling at approximately 67% Vo(2 max) with water (WAT) or CHO ingestion (1.5 g of glucose/min). Substrate oxidation and kinetics were quantified during exercise using indirect calorimetry and stable isotope techniques ([(13)C]glucose ingestion, [6,6-(2)H(2)]glucose, and [(2)H(5)]glycerol infusion). In both sexes, CHO ingestion significantly increased the rates of appearance (R(a)) and disappearance (R(d)) of glucose during exercise compared with WAT ingestion [males: WAT, approximately 28-29 micromol x kg lean body mass (LBM)(-1) x min(-1); CHO, approximately 53 micromol x kg LBM(-1) x min(-1); females: WAT, approximately 28-29 micromol x kg LBM(-1) x min(-1); CHO, approximately 61 micromol x kg LBM(-1) x min(-1); main effect of trial, P < 0.05]. The contribution of plasma glucose oxidation to the energy yield was significantly increased with CHO ingestion in both sexes (from approximately 10% to approximately 20% of energy expenditure; main effect of trial, P < 0.05). Liver-derived glucose oxidation was reduced, although the rate of muscle glycogen oxidation was unaffected with CHO ingestion (males: WAT, 108 +/- 12 micromol x kg LBM(-1) x min(-1); CHO, 108 +/- 11 micromol x kg LBM(-1) x min(-1); females: WAT, 89 +/- 10 micromol x kg LBM(-1) x min(-1); CHO, 93 +/- 11 micromol x kg LBM(-1) x min(-1)). CHO ingestion reduced fat oxidation and lipolytic rate (R(a) glycerol) to a similar extent in both sexes. Finally, ingested CHO was oxidized at similar rates in men and women during exercise (peak rates of 0.70 +/- 0.08 and 0.65 +/- 0.06 g/min, respectively). The present investigation suggests that the metabolic response to CHO ingestion during exercise is largely similar in men and women.     

Roles of nitric oxide and angiotensin II in the impaired baroreflex gain of pregnancy

The present study tested the hypothesis that nitric oxide (NO) contributes to impaired baroreflex gain of pregnancy and that this action is enhanced by angiotensin II. To test these hypotheses, we quantified baroreflex control of heart rate in nonpregnant and pregnant conscious rabbits before and after: 1) blockade of NO synthase (NOS) with Nomega-nitro-L-arginine (20 mg/kg iv); 2) blockade of the angiotensin II AT1 receptor with L-158,809 (5 microg x kg(-1) x min(-1) iv); 3) infusion of angiotensin II (1 ng x kg(-1) x min(-1) nonpregnant, 1.6-4 ng x kg(-1) x min(-1) pregnant iv); 4) combined blockade of angiotensin II AT(1) receptors and NOS; and 5) combined infusion of angiotensin II and blockade of NOS. To determine the potential role of brain neuronal NOS (nNOS), mRNA and protein levels were measured in the paraventricular nucleus, nucleus of the solitary tract, caudal ventrolateral medulla, and rostral ventrolateral medulla in pregnant and nonpregnant rabbits. The decrease in baroreflex gain observed in pregnant rabbits (from 23.3 +/- 3.6 to 7.1 +/- 0.9 beats x min(-1) x mmHg(-1), P < 0.05) was not reversed by NOS blockade (to 8.3 +/- 2.5 beats x min(-1) x mmHg(-1)), angiotensin II blockade (to 5.0 +/- 1.1 beats x min(-1) x mmHg(-1)), or combined blockade (to 12.3 +/- 4.8 beats x min(-1) x mmHg(-1)). Angiotensin II infusion with (to 5.7 +/- 1.0 beats x min(-1) x mmHg(-1)) or without (to 8.4 +/- 2.4 beats x min(-1) x mmHg(-1)) NOS blockade also failed to improve baroreflex gain in pregnant or nonpregnant rabbits. In addition, nNOS mRNA and protein levels in cardiovascular brain regions were not different between nonpregnant and pregnant rabbits. Therefore, we conclude that NO, either alone or via an interaction with angiotensin II, is not responsible for decrease in baroreflex gain during pregnancy.