Spatio-temporal dynamics in maturation and spawning of Patagonian toothfish Dissostichus eleginoides on the sub-Antarctic Kerguelen Plateau.

This study investigated maturation and spawning of Patagonian toothfish Dissostichus eleginoides in the Heard Island and McDonald Islands (HIMI) fishery on the Kerguelen Plateau in the Indian Sector of the Southern Ocean based on gonads and otoliths collected between 2004 and 2015 and using histological analyses and calibration of macroscopic staging criteria. Dissostichus eleginoides at HIMI spawn throughout the austral late autumn and winter months of May-August and spawning activity is concentrated on slopes along the west and south of the plateau around HIMI at depths of 1500-1900 m. Comparison between histological analyses and macroscopic gonad staging indicated that many fish that had spawned, as indicated by the presence of post-ovulatory follicles, returned to a resting stage which was macroscopically indistinguishable from maturing fish. Furthermore, the occurrence of females of all size classes with low gonado-somatic index and low macroscopic gonad stage during the spawning season suggested that a proportion of mature females did not spawn every year. Age-at-maturity estimates, based on the assumption that fish of macroscopic stages ≥2 were mature, decreased between the 2004-2009 and 2010-2015 periods for both sexes. The magnitude of this temporal variation in age at maturity, however, varied between gear types and fishing depths and variable sampling regimes probably influenced these variations. This study highlights the importance of correct interpretation of macroscopic gonad stages and understanding the influence of fishery operations on estimations of life-history parameters.

Targeted alpha(1A)-adrenergic receptor overexpression induces enhanced cardiac contractility but not hypertrophy.

Activation of the alpha(1A)-adrenergic receptor (alpha(1A)-AR)/Gq pathway has been implicated as a critical trigger for the development of cardiac hypertrophy. However, direct evidence from in vivo studies is still lacking. To address this issue, transgenic mice with cardiac-targeted overexpression of the alpha(1A)-AR (4- to 170-fold) were generated, using the rodent alpha-myosin heavy chain promoter. Heterozygous animals displayed marked enhancement of cardiac contractility, evident from increases in dP/dt(max) (80%, P<0.0001), dP/dt(max)/LVP(inst) (76%, P<0.001), dP/dt(max):dP/dt(min) (104%, P<0.0001), and fractional shortening (33%, P<0.05). Moreover, changes in the dP/dt(max)-end-diastolic volume relationship provided load-independent evidence of a primary increase in contractility. Blood pressure and heart rate were largely unchanged, and there was a small increase in (-)norepinephrine-stimulated, but not basal, phospholipase C activity. Increased contractility was directly related to the level of receptor overexpression and could be completely reversed by acute alpha(1A)- but not beta-AR blockade. Despite the robust changes in contractility, transgenic animals displayed no morphological, histological, or echocardiographic evidence of left ventricular hypertrophy. In addition, apart from an increase in atrial natriuretic factor mRNA, expression of other hypertrophy-associated genes was unchanged. To our knowledge, these data provide the first in vivo evidence for an inotropic action of the alpha(1A)-AR.

An NMR investigation of electron transfer in the copper-protein, plastocyanin.

1. The proton NMR spectra of oxidised and reduced French bean plastocyanin have been recorded on a 270 MHz pulsed spctrometer. 2. The spectrum of a mixture containing the protein in the paramagnetic Cu(II) and diamagnetic Cu(I) states is a superposition of the separate spectra. When ferrirate spectra. 3. The results show that self-exchange between Cu(II)- and Cu(I)-plastocyanin is slow on the NMR time scale (kex less than 2-10(4) M-1-s-1 at 50 degrees C), and that electron transfer in the presence of ferricyanide is rapid (k greater than 1-10(5) M-1-s-1).

Inositol phosphates in the heart: controversy and consensus.

Numerous studies have addressed various aspects of inositol phosphate release and metabolism in myocardial preparations, and many different viewpoints have been expressed. The various results and interpretations presented often appear confusing and extracting a consensus view can be difficult. The differences often derive from the differing cardiac preparations used, especially isolated cells versus intact tissue. Despite these problems there are aspects where consensus prevails. Both the metabolism and the functional activity of inositol phosphates in heart appear to differ from those previously described in non-excitable cells. Inositol phosphates do not appear to be of major importance in the control of cardiac function under physiological conditions but may well have greater influence under pathological conditions such as myocardial ischaemia and reperfusion. Hopefully, the near future will see remaining controversies resolved.

Beta(2)-adrenergic receptor overexpression driven by alpha-MHC promoter is downregulated in hypertrophied and failing myocardium.

OBJECTIVE: The alpha-myosin heavy chain (alpha-MHC) promoter is frequently used to direct cardiac specific transgene expression. We studied whether transgene expression controlled by this promoter was altered under conditions of cardiac hypertrophy and failure. METHODS: Transgenic (TG) mice overexpressing human beta(2)-adrenergic receptors (beta(2)AR) and wild type (WT) controls were subjected to thoracic aortic constriction (TAC) or sham operation and studied at 1, 3 and 8 weeks after surgery. RESULTS: Sham operated TG mice had higher heart rates and left ventricular (LV) contractility than WT (all P<0.01), demonstrating enhanced betaAR activation. TAC at 1, 3 and 8 weeks produced progressive LV hypertrophy which was similar between WT and TG mice. Evidence of heart failure was more marked in TG mice with a greater increase in weights of the right ventricle and lungs and a higher prevalence of atrial thrombus (P<0.05 in each case). In hypertrophied TG hearts, endogenous alpha-MHC mRNA transcripts in LV were maintained at 1 and 3 weeks, but were reduced by approximately 40% relative to the sham-operated group at 8 weeks after TAC. Transgene expression, measured as human beta(2)AR mRNA, was reduced by 45% at 1 and 3 weeks and by 70% at 8 weeks after TAC. beta(2)AR binding sites were reduced by 35, 47 and 65%, respectively, at 1, 3 and 8 weeks. CONCLUSION: Cardiac hypertrophy and failure cause downregulation of the endogenous alpha-MHC as well as cardiac specific overexpression of the transgene directed by an alpha-MHC promoter.

Age-dependent cardiomyopathy and heart failure phenotype in mice overexpressing beta(2)-adrenergic receptors in the heart.

OBJECTIVE: To explore long-term cardiac phenotype in transgenic (TG) mice with 300-fold overexpression of beta(2)-adrenergic receptors (AR). METHODS: Echocardiography was performed serially on a cohort of wild-type and TG mice (n=26 each) between 4 and 15 months of age. Survival was monitored and autopsy and histological examinations were performed. RESULTS: Heart rate was higher in TG than in wild-type mice throughout the study period. The left ventricular dimensions and fractional shortening were similar between TG and wild-type groups during 4-6 months. Starting at 9 months, however, TG mice showed progressive reduction in fractional shortening and systolic wall thickening, and increase in left ventricular dimensions and left ventricular mass, indicating onset of heart failure, left ventricular hypertrophy and remodeling. Abnormal waveforms in the electrocardiogram and episodes of ventricular ectopic beats were also observed in TG mice. Death of TG mice started at 8.5 months, and the cumulative mortality was 81% by 15 months (P<0.0001 vs. 4% in wild-type mice). The majority of deaths were due to severe heart failure, indicated by cardiac dilatation, lung congestion, pleural effusion and atrial thrombus. Left ventricular sections showed widespread interstitial fibrosis, loss of myocytes and myocyte hypertrophy in TG mice. CONCLUSIONS: A high level of beta(2)AR overexpression results in cardiomyopathy and heart failure. The onset was slower and the expression levels of receptors required are much higher than previously described for the beta(1)AR overexpression.

Inhibition of adenylate cyclase in rat adrenal glomerulosa cells by angiotensin II.

Adenylate cyclase in homogenates of rat adrenal glomerulosa cells was inhibited in a concentration dependent manner by angiotensin II (AII). The maximum inhibition of basal activity was 34.5 +/- 2.7% and the EC50 value for AII was 1.1 +/- 0.4 nM (n = 6). Similar maximum inhibitions were produced by the precursor decapeptide, AI, and the heptapeptide, AIII [(des asp) AII]. The EC50 values for these two peptides were respectively 72 +/- 8 nM and 25 +/- 5 nM (n = 6). The antagonist compound (1-sarcosine, 8-isoleucine)-AII, reversed the effect of AII. Inhibition of the adrenal enzyme required guanosine triphosphate and monovalent cations as has been described for adenylate cyclase inhibition in other tissues. Maximum inhibition was observed at 10(-5) M guanosine triphosphate and 150 mM Na+ or Li+ ion. Both basal adenylate cyclase activity and activity stimulated by adrenocorticotrophic hormone were inhibited. These results demonstrate the presence in rat adrenal glomerulosa cells of angiotensin receptors coupled to adenylate cyclase inhibition and show that their properties are similar to those of adrenal angiotensin receptors described previously.

Renal proximal tubular alpha-adrenergic receptors oppose urinary 3,5'-cyclic adenosine monophosphate response to parathyroid hormone in vivo.

In both man and rat, urinary cAMP (U cAMP) level increases in response to PTH. The increased cAMP arises largely by secretion from the proximal tubule where cAMP synthesis is stimulated by PTH through adenylate cyclase-coupled receptors. We have previously demonstrated alpha 2-adrenergic receptors which inhibit PTH-stimulated adenylate cyclase in rat renal cortex membranes in vitro. In the present study, the effects of alpha-adrenergic agonists and antagonists on the U cAMP response to PTH were investigated in anesthetized rats in vivo. Injection of PTH (15 U/kg iv) produced an increase in U cAMP from 1.7 +/- 0.3 to 7.4 +/- 0.7 nmol cAMP/mumol creatinine (n = 6), (P less than 0.001). This rise was largely due to an increase in nephrogenous cAMP which increased 10-fold. Infusion of the alpha 2-adrenergic agonist clonidine at 1 microgram/kg X min caused a decrease in the cAMP response to PTH to 3.6 +/- 0.5 nmol cAMP/mumol creatinine (n = 12) (P less than 0.001). Infusion of the alpha 2-selective catecholamine alpha-methylnorepinephrine (1 microgram/kg X min) caused a similar reduction in U cAMP response to that observed with clonidine. The alpha-adrenergic antagonist phentolamine (100 micrograms/kg X min) reversed the effects of clonidine and, when administered in the absence of alpha-agonists, caused an increased cAMP response to PTH. These results demonstrate the presence of alpha-receptors in the rat proximal convoluted tubule which oppose the actions of PTH in vivo.

Inhibition of adenylate cyclase by angiotensin II in rat renal cortex.

Adenylate cyclase of rat renal cortex was inhibited by angiotensin II (AII). Inhibition required Na+ (100-200 mM) and GTP (10(-8)-10(-4) M) and was opposed by the receptor antagonist [1-sarcosine, 8-isoleucine]AII. The EC50 value (+/- SE)for inhibition by AII was 3.7 +/- 1.2 nM, and the maximum inhibition (+/- SE) was 23 +/- 3%. Inhibition was specific for AII, since both AI and AIII, at concentrations up to 1 microM, were ineffective in producing inhibition. The maximum decrease (+/- SE) in adenylate cyclase activity was from 2.45 +/- 0.08 to 1.78 +/- 0.1 pmol.min/mg protein. A similar absolute decrease was observed when adenylate cyclase was stimulated by calcitonin, vasopressin, or isoproterenol. The inhibition of PTH-stimulated activity [16.7 +/- 0.5 (+/- SE) to 12.2 +/- 0.7 pmol.min/mg protein) was significantly greater than the inhibition of basal activity. Therefore, at least some of the inhibitory angiotensin receptors are coupled to adenylate cyclase molecules which also coupled to receptors for PTH.

Adenylate cyclase inhibition is not involved in the adrenal steroidogenic response to angiotensin II.

Angiotensin II (AII) receptors in adrenal glomerulosa cells are coupled to adenylate cyclase inhibition. We investigated the importance of cyclase inhibition in adrenal steroidogenesis by treating adrenal glomerulosa cells with the toxin of Bordetella pertussis (20 ng/ml) for 3 and 18 h. This treatment prevented inhibition of forskolin-stimulated adenylate cyclase by AII. However, the aldosterone response to AII was not altered by toxin treatment. These results strongly suggest that adenylate cyclase inhibition is not directly involved in mediating the adrenal actions of AII. In addition, ACTH-induced steroidogenesis also was unaffected by toxin treatment demonstrating that cyclase inhibition is not involved in suppressing steroidogenesis via the cAMP pathway.

Vasopressin stimulates phosphatidylinositol turnover and aldosterone synthesis in rat adrenal glomerulosa cells: comparison with angiotensin II.

Vasopressin (VP) action was identified in rat adrenal glomerulosa cells measuring the VP stimulation of phosphatidylinositol breakdown to inositol-1-phosphate (IP). The pKb value (negative log of EC50) for arginine VP (AVP) was 9.1 +/- 0.4 (n = 6). The V2-selective agonist 1-deamino-8-D-arginine VP did not stimulate IP accumulation. Furthermore, the V1-selective antagonist [1-(beta-mercapto-beta, beta-cyclopentamethylenepropionic acid) 2-(O-methyl) tyrosine]AVP (10(-7)M) prevented the stimulation by VP. Thus, the VP receptors in adrenal glomerulosa cells appeared to be V1-type similar to those found in liver and vasculature and different from those in kidney. Angiotensin II (AII) also stimulated accumulation of IP but the maximum stimulation achieved was much greater than with VP. In the case of AII, stimulation of phosphatidylinositol breakdown is thought to be the initiating event in the stimulation of aldosterone synthesis. In agreement with this, both VP and AII stimulated aldosterone synthesis, the maximum AII stimulation (6.5- +/- 1.3-fold, n = 7) being much greater than the VP stimulation (1.7 +/- 0.33-fold, n = 7).

Angiotensin II-stimulated phosphatidylinositol turnover in rat adrenal glomerulosa cells has a complex dependence on calcium.

The stimulation of phosphatidylinositol (PI) turnover by angiotensin II in rat adrenal glomerulosa cells has been studied in detail and shown to have a complex dependence on Ca2+. After the addition of angiotensin II, inositol monophosphate, inositol bisphosphate, and inositol trisphosphate increased rapidly and transiently. The transient increase was followed by a slower sustained rise, which continued for up to 30 min. Inositol phosphate accumulation during the sustained phase was decreased when experiments were performed in Ca2+-free medium. The initial transient response was not altered. Addition of the Ca2+ ionophore A23187 enhanced the angiotensin II response at 20 min, but not the 15 sec response. The sustained response, but not the transient response, was attenuated by the Ca2+ channel blocker nifedipine, indicating that the effect of Ca2+ required uptake through voltage-dependent Ca2+ channels. Subsequent studies showed that cAMP decreased inositol phosphate accumulation at 20 min while having no effect at 15 sec. Also, incubation with phorbol 12-myristate 13-acetate produced a more effective inhibition of the sustained response than of the initial transient response. However, while the transient and sustained phases of PI turnover were differently affected by Ca2+ and inhibitory compounds, the profiles of inositol phosphates generated were similar. At both 15 sec and 20 min inositol-(1,4,5) trisphosphate was detected, indicating sustained cleavage of PI-(4,5) bisphosphate. Taken together, the results suggest that while the initial PI turnover response is independent of Ca2+ and presumably initiates the rise in cytosolic Ca2+, sustained response requires entry of Ca2+ to maintain elevated cytosolic Ca2+ concentrations. Thus, while the increase in cytosolic Ca2+ may have a direct role in the stimulation of aldosterone synthesis, it is also required to sustain the PI turnover response to angiotensin II.

Properties of [3H]1-desamino-8-D-arginine vasopressin as a radioligand for vasopressin V2-receptors in rat kidney.

[3H]1-Desamino-8-D-arginine vasopressin [3H] DDAVP was assessed as a radioligand for vasopressin V2-receptors by studying its membrane-binding characteristics and in vitro autoradiographic localization in rat kidney, a rich source of V2-receptors. [3H]DDAVP bound specifically to a single class of high affinity, low capacity sites in rat medullopapillary membranes. Specific [3H]DDAVP binding at 25 C reached equilibrium after 2 h of incubation and was saturable and linear with protein concentration up to 2.2 mg/ml protein. Saturation analysis gave an equilibrium dissociation constant (Kd) of 0.76 nM. Displacement of [3H]DDAVP binding by unlabeled arginine vasopressin (AVP) and related analogs gave the following order of potency, consistent with that expected for a V2-receptor: DDAVP approximately equal to AVP approximately equal to 1-desamino-AVP greater than lysine vasopressin greater than oxytocin greater than [1-(beta-mercapto-beta, beta-cyclopentamethylene-propionic acid, 2-(O-methyl)tyrosine]AVP. The C-terminal metabolites of AVP, (pGlu4Cyt6)AVP-(4-9), and (pGlu4Cyt6)AVP-(4-8) did not displace [3H]DDAVP binding. No degradation of [3H] DDAVP during incubation could be detected by HPLC analysis. In vitro autoradiography of [3H]DDAVP binding to rat kidney sections showed a very dense localization of displaceable binding over inner and outer medulla, with a much lower density in cortex, consistent with the known major localization of V2-receptors on renal collecting tubules. These studies suggest that [3H]DDAVP is a suitable radioligand for labeling V2-receptors and may be useful in the characterization of vasopressin receptor subtypes in a variety of tissues and in purification of the V2-receptor.

Changes in tissue alpha- and beta-adrenergic receptors in renal hypertension in the rat.

Myocardial membranes prepared from renal hypertensive rats contained reduced concentrations of both alpha- and beta-adrenergic receptors. The decrease in alpha-receptor concentration measured by [3H]-dihydroergocryptine binding was from 80 +/- 6 (SEM) to 52 +/- 2 fmol/mg. Beta-receptor concentration measured by 125I-iodohydroxybenzylpindolol binding also decreased by about half from 80 +/- 16 to 41 +/- 9 fmol/mg. The affinities of the receptors were unchanged. There was no change in either concentration or affinity of beta receptors in membranes prepared from the lungs or kidneys of these hypertensive rats. There results demonstrate that the observed receptor changes are tissue-specific. Cardiac adrenergic receptor alterations are therefore not part of a generalized adrenergic receptor decrease associated with elevated circulating plasma catecholamine concentrations, but probably reflect a specific increase in cardiac sympathetic drive.

Sympathetic activity and cardiac adrenergic receptors in one-kidney, one clip hypertension in rats.

The activity of the sympathetic nervous system, as measured by levels of plasma and cardiac catecholamines and catecholamine metabolites and the function of cardiac alpha- and beta-adrenergic receptors, was evaluated at 3 days and 4 weeks after induction of one-kidney, one clip hypertension (1K1C) in the rat. At 3 days, the plasma level of norepinephrine (NE) was lower in the 1K1C group than the control group (p less than 0.01), whereas epinephrine (E) and the metabolites dihydroxymandelic acid (DOMA), dihydroxyphenylglycol (DOPEG), and normetanephrine (NMN) were similar in both groups. In addition, cardiac content of catecholamines, their metabolites, and adrenergic receptors were similar in both groups. At 4 weeks, plasma levels of NE and DOPEG were lower (p less than 0.01), whereas levels of DOMA and NMN were higher (p less than 0.02 and p less than 0.001, respectively) in the 1K1C group than the control group. Cardiac content of NE (p less than 0.01), and DOPEG (p less than 0.05) was significantly lower, whereas DOMA and NMN were significantly higher (p less than 0.01) in the 1K1C group as compared to controls. In addition, cardiac density of both alpha- and beta-adrenergic receptors was reduced in the 1K1C group, whereas receptor affinities were unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)

Specific changes in hypothalamic alpha-adrenoceptors in young spontaneously hypertensive rats.

Changes in the activity of hypothalamic and brain-stem adrenergic neurons have been reported in young spontaneously hypertensive rats (SHR) prior to the development of hypertension. We have measured central alpha- and beta-adrenoceptor concentrations in 4-week-old SHR and Wistar-Kyoto (WKY) controls by direct radioligand binding studies using [3H]prazosin (alpha 1), [3H]clonidine (alpha 2), and [125I]iodohydroxybenzlpindolol (beta). The concentration of alpha 2-adrenoceptors was significantly elevated in the hypothalamus of the SHR, 156.9 +/- 10.4 compared with WKY, 119.4 +/- 10.0 fmole/mg protein (n = 7, mean +/- SEM, p less than 0.0125). Alpha 2-adrenoceptor concentrations in both the brain stem and cerebral cortex were similar in the two groups of animals. The increase in hypothalamic adrenoceptors was specific for alpha 2-adrenoceptors, since similar concentrations of alpha 2- and beta-adrenoceptors were found in this region.

Right atrial dilatation increases inositol-(1,4,5)trisphosphate accumulation. Implications for the control of atrial natriuretic peptide release.

Stretching the right atrium of isolated perfused [3H]inositol-labelled rat hearts was shown to stimulate the phosphatidyl-inositol turnover pathway as demonstrated by the accumulation of [3H]inositol-(1,4,5)trisphosphate and its degradation products. Stimulation was detectable after 1 min with larger increases observed after 10 or 20 min. These findings demonstrate that the myocardium can respond to dilatation by an activation of the phosphatidylinositol turnover pathway. Such a mechanism has implications for the release of atrial natriuretic peptide following right atrial distention.

Isolation, amino acid sequence and action of guinea-pig ACTH on aldosterone production by glomerulosa cells.

Though minor sequence differences between-species have been reported for adrenocorticotrophin, ACTH(1-39), the steroidogenic moiety ACTH(1-24) has appeared invariant in mammals. We here report the isolation, purification and amino acid sequencing of guinea-pig (GP) ACTH in which Pro24 is replaced by Ala24, and the demonstration that GP-ACTH stimulates aldosterone production to maximal levels well above those seen with human ACTH(1-39) or Synacthen, ACTH(1-24) amide, the synthetic ACTH fragment widely used for diagnostic and therapeutic purposes.

Adrenocorticotropic hormone inhibits angiotensin II-stimulated inositol phosphate accumulation in rat adrenal glomerulosa cells.

Rat adrenal glomerulosa cells labelled for 18 h with [3H]inositol responded to angiotensin II with a dose-dependent stimulation of the accumulation of inositol monophosphate, inositol bisphosphate and inositol trisphosphate. Addition of adrenocorticotropic hormone (ACTH) (10(-7)M) reduced the maximum responses without altering the EC50 values for angiotensin II. Thus, ACTH acted as a non-competitive inhibitor with respect to angiotensin II. No inhibition was observed in cells labelled for 2 h with [3H]inositol. Detailed examination of the inhibition showed that ACTH(1-24) was the most potent inhibitor, with ACTH(1-39) being 10-fold less potent. A mixture of alpha-melanocyte-stimulating hormone (alpha-MSH) (ACTH(1-13] and corticotropin-like intermediate lobe peptide (ACTH(18-39] was similarly inactive. ACTH(5-24) did not produce detectable inhibition. In terms of specificity, the receptor mediating ACTH inhibition of phosphatidylinositol turnover was similar to the receptor which mediated stimulation of aldosterone synthesis. Inhibition by ACTH was additive with inhibition produced by dibutyryl cAMP demonstrating that it was not mediated by rises in intracellular cAMP. ACTH inhibition also was additive with inhibition by the calcium channel blocker, nifedipine. These results demonstrate an interaction between ACTH receptors and angiotensin II receptors in adrenal glomerulosa cells at the level of their receptor-second messenger pathways.

Postnatal development of rat alpha- and beta-adrenergic receptors: A comparison between tissues.

The rates of development of rat kidney α- and ß-adrenoceptors were compared with those of heart and lung adrenoceptors in the same animals by direct binding studies using [(3)H]WB4101 (α1), [(3)H]yohimbine (α2) and [(125)I]HYP (ß). Kidney α1 and ß-adrenoceptors had reached adult concentrations 7 days after birth, while the α2-adrenoceptor concentration plateaued at 21 days. Lung ß-adrenoceptor concentration was stable initially, then increased rapidly to adult levels by 18 days. In contrast, heart α1-and ß-adrenoceptor concentrations were at mature levels at birth. In all tissues studied the increase in noradrenaline concentration was slower than the increases in adrenoceptor concentrations.