Enantioselective total synthesis and biological evaluation of (-)-solanacol.

An enantioselective synthesis of the phenyl ring-containing strioglactone, (-)-solanocol, is described. Application of a Dynamic Kinetic Resolution (DKR) in the stereo-defining step enabled a step-economical synthesis to be achieved, and allowed access to natural and non-natural enantiomers with equal facility. Results of seed germination assays and Differential Scanning Fluorimetry (DSF) measurements with the known strigolactone receptor protein, Decreased Apical Dominance 2 (DAD2), are reported.

Pharmacokinetic and pharmacodynamic properties of SOL1: a novel dual inhibitor of neutral endopeptidase and endothelin converting enzyme.

AIMS: The pharmacological profile of the novel putative neutral endopeptidase (NEP) and endothelin converting enzyme (ECE) inhibitor SOL1 was examined. MAIN METHODS: The enzyme inhibitory profile of SOL1 was established in vitro. The pharmacokinetic and pharmacodynamic profile was determined in rodents in vivo. KEY FINDINGS: In vitro, at neutral pH, 10 µM SOL1 inhibited NEP-1, NEP-2, and ECE-1 by 99%, 94% and 75%, respectively. The IC(50)s were 25, 25 and 3200 nmol/L, respectively. In anesthetized rats, SOL1 inhibited blood pressure (BP) responses to big-ET-1 and ET-1(1-31) with ED(50)s of 1.9 and 0.03 mg/kg, corresponding to plasma EC(50)s of 4.6 and 0.1 µmol/L, respectively. Pharmacokinetics of SOL1 were examined after single injections in mice and rats. In these species, the estimated clearance of SOL1 varied between 5 and 9 ml/kg.min and T(1/2) between 20 and 60 min. Steady state kinetics of SOL1 were examined after continuous s.c. infusions of SOL1 for 3 weeks at 50mg/kg.day in DOCA-salt hypertensive rats. This treatment lowered BP by 22 mmHg. Steady state concentrations of SOL1 in plasma were 3.9 µmol/L. In heart, lung, and kidney the concentrations of SOL1 were 0.4, 1.8, and 20.5 µmol/kg, respectively. About 63% of the daily dose was retrieved unaltered in the urine. SIGNIFICANCE: These data indicate that SOL1 is primarily a NEP inhibitor in vitro as well as in vivo. Given the preferential renal accumulation and renal clearance of SOL1 additional ECE-1 inhibition in the kidney may have contributed to its chronic BP lowering effects in the DOCA-salt hypertensive rat model.

Impaired cardiac functional reserve in type 2 diabetic db/db mice is associated with metabolic, but not structural, remodelling.

AIM: To identify the initial alterations in myocardial tissue associated with the early signs of diabetic cardiac haemodynamic dysfunction, we monitored changes in cardiac function, structural remodelling and gene expression in hearts of type 2 diabetic db/db mice. METHODS: Cardiac dimensions and function were determined echocardiographically at 8, 12, 16 and 18 weeks of age. Left ventricular pressure characteristics were measured at 18 weeks under baseline conditions and upon dobutamine infusion. RESULTS: The db/db mice were severely diabetic already at 8 weeks after birth, showing elevated fasting blood glucose levels and albuminuria. Nevertheless, echocardiography revealed no significant changes in cardiac function up to 18 weeks of age. At 18 weeks of age, left ventricular pressure characteristics were not significantly different at baseline between diabetic and control mice. However, dobutamine stress test revealed significantly attenuated cardiac inotropic and lusitropic responses in db/db mice. Post-mortem cardiac tissue analyses showed minor structural remodelling and no significant changes in gene expression levels of the sarcoplasmic reticulum calcium ATPase (SERCA2a) or beta1-adrenoceptor (beta1-AR). Moreover, the phosphorylation state of known contractile protein targets of protein kinase A (PKA) was not altered, indicating unaffected cardiac beta-adrenergic signalling activity in diabetic animals. By contrast, the substantially increased expression of uncoupling protein-3 (UCP3) and angiopoietin-like-4 (Angptl4), along with decreased phosphorylation of AMP-activated protein kinase (AMPK) in the diabetic heart, is indicative of marked changes in cardiac metabolism. CONCLUSION: db/db mice show impaired cardiac functional reserve capacity during maximal beta-adrenergic stimulation which is associated with unfavourable changes in cardiac energy metabolism.

Acute autonomic effects of vitamins and fats in male smokers.

BACKGROUND/OBJECTIVES: Vitamins can help improve cardiovascular control. In contrast, smoking works in the opposite fashion, reducing the baroreflex control of heart rate (HR) possibly via oxidative stress. High-fat challenges also impair cardiovascular regulation. Whether vitamins have acute beneficial effects on the baroreflex control of HR in smokers is unclear. SUBJECTS/METHODS: A randomized, placebo-controlled crossover study in 30 male smokers (34.2+/-6.9 years). Interventions were: (1) moderate (vitamin C (300 mg) and E (75 IU) and folic acid (1 mg)); (2) high doses of vitamins (vitamin C (2 g) and E (800 IU), and folic acid (5 mg)); or, (3) placebo. Vitamins were ingested with cream (a high-fat challenge) or milk (low-fat control). Four hours later, blood was withdrawn and radial pulse wave forms recorded via tonometry. Spontaneous beat-to-beat variations in HR and systolic blood pressure (SBP) were analysed by spectral analysis techniques and sympathovagal control of HR and baroreflex sensitivity (BRS) were assessed. RESULTS: High doses of vitamins increased plasma vitamin C, E and folic acid levels (P<0.05) with no change in SBP, HR or BRS (P>0.05, analysis of variance). Plasma vitamin levels did not correlate with any cardiovascular parameters. Moderate vitamins increased the vagal control of HR (+23%; P<0.05) and cream led to small increases (P<0.05) in SBP (+2 mm Hg) and HR (+2 beats min(-1)) with no change in BRS. CONCLUSIONS: In male smokers, circulating antioxidants had no effect on BRS and minor effects on the cardiovascular system were seen following acute fat and vitamin ingestion.

Renal medullary effects of transient prehypertensive treatment in young spontaneously hypertensive rats.

AIM: Transient angiotensin II receptor blockade (ARB) leads to prolonged blood pressure (BP) lowering, but the underlying mechanism remains uncertain. Long-term BP control is regulated by the medullary microcirculation with the pericyte as contractile cell. We hypothesize that the prolonged BP effect is caused by increased medullary blood flow (MBF) associated with structural alterations based on reduced medullary pericyte number. METHODS: Four-week-old spontaneously hypertensive rats (SHR) were treated for 4 weeks with losartan (SHR-Los: 20 mg kg(-1) day(-1)), hydralazine (SHR-Hyd: 15 mg kg(-1) day(-1)), losartan and pan-caspase inhibitor zVAD (SHR-Los + 1 mg kg(-1) day(-1) zVAD), losartan and glycogen synthase kinase-3beta (GSK) inhibitor valproate (SHR-Los + 10 mg kg(-1) day(-1) Val) or placebo. BP, MBF and pericyte number were determined under and after treatment (8 and 12 weeks). Apoptotic pericytes were determined with alpha-actin and TUNEL double staining. Sodium concentration was determined in renal medulla and urine. RESULTS: Antihypertensive treatment equipotently reduced BP at 8 weeks of age. After drug withdrawal (12 weeks of age) BP reduction was restricted to SHR-Los (SHR-Los: 153 +/- 5, SHR-Hyd: 177 +/- 2, SHR: 184 +/- 3 mmHg). Simultaneously, MBF was increased and pericyte number reduced, while medullary and urinary sodium concentration increased. Transient ARB in combination with zVAD or valproate resulted in more medullary pericytes and higher BP (SHR-Los/zVAD: 164 +/- 7; SHR-Los/Val: 168 +/- 6 mmHg) compared with transient ARB alone. CONCLUSION: After drug withdrawal, transient ARB leads to increased MBF and is associated with a reduction in medullary pericytes. This may be associated with pericyte apoptosis as anti-apoptosis during transient ARB increases pericyte number and BP.

Oestrogen modulates cardiac ischaemic remodelling through oestrogen receptor-specific mechanisms.

AIM: Observational and clinical studies suggest different responses upon sex hormone replacement therapy in ischaemic heart disease. Few studies, however, have examined the impact of oestrogen receptor-dependent mechanisms on the extent of injury after myocardial infarction (MI). Therefore, we set out to evaluate the effect of oestrogen (E2) replacement on infarct size and remodelling, and the respective role of the oestrogen receptors (ER)alpha and -beta in this process, using ERalpha- and ERbeta-deficient mice. METHODS: Wild type (WT) (ERalpha(+/+) and ERbeta(+/+)), ERalpha-deficient (ERalpha(-/-)) and ERbeta-deficient (ERbeta(-/-)) mice were ovariectomized and subsequently supplemented with E2 or placebo using subcutaneous 60-day release pellets. MI was induced by left coronary artery ligation. Two weeks following MI, haemodynamic function was assessed and infarct size was determined. RESULTS: There was no significant difference in infarct size between E2- or placebo-treated WT (ERalpha(+/+) and ERbeta(+/+)) mice. Surprisingly, E2 treatment did result in smaller infarct sizes in ERalpha(-/-) mice, but increased the infarct size in ERbeta(-/-) mice. Increase of the left ventricular mass post-MI was significantly larger in the E2-treated ERalpha(-/-) animals compared with placebo-treated animals. E2 treatment also significantly increased post-MI mortality in ERalpha(+/+), ERbeta(+/+) and ERalpha(-/-) animals, but not in ERbeta(-/-) mice. CONCLUSIONS: Although E2 modulates the infarct size in ERalpha(-/-), it also appears to be responsible for the higher mortality following MI. ERbeta appears to be the receptor involved in the modulating effects of E2 in the infarcted heart.

Specific and sustained down-regulation of genes involved in fatty acid metabolism is not a hallmark of progression to cardiac failure in mice.

Preferential and specific down-regulation of genes involved in fatty acid (FA) uptake and metabolism is considered a hallmark of severe hypertrophic remodeling and progression to cardiac failure. Therefore, we investigated the time course of changes in cardiac metabolic gene expression (1) in mice subjected to regional myocardial infarction (MI) for 4 days, 1 month, or 3 months and (2) in mice overexpressing calcineurin (Cn) which initially develop concentric hypertrophy progressing after the age of 4 weeks to dilated cardiomyopathy and failure. In both models, hypertrophy was characterized by increased expression of beta-myosin heavy chain protein and atrial natriuretic factor mRNA, indicative of marked structural remodeling. Fractional shortening progressively decreased from 31% to 15.1% and 3.7% 1 and 3 months after MI, respectively. One month post-MI, the expression of several metabolic genes, i.e., acyl-CoA synthetase (-50%), muscle-type carnitine palmitoyl transferase 1 (-37%) and citrate synthase (-28%), was significantly reduced in the surviving myocardium. Despite overt signs of cardiac failure 3 months post-MI, the expression of these genes had returned to normal levels. In hearts of both 4- and 6-week-old Cn mice, genes involved in both FA and glucose metabolism and mitochondrial citrate synthase were down-regulated, reflecting an overall decline in metabolic gene expression, rather than a specific and preferential down-regulation of genes involved in FA uptake and metabolism. These findings challenge the concept that specific and sustained down-regulation of genes involved in FA uptake and metabolism represents a hallmark of the development of cardiac hypertrophy and progression to failure.

Chronic moderate hypoxia during in ovo development alters arterial reactivity in chickens.

We previously observed arterial sympathetic hyperinnervation and endothelial dysfunction in the chicken embryo after exposure to chronic hypoxia. We now investigate whether changes in arterial properties could also be observed at 14-15 weeks of life. Eggs of White Leghorn chicken were incubated under normoxic or moderately hypoxic (15% O2 from days 6-19 of a 21-day incubation) conditions. Experiments were performed at 14-15 weeks of life under standard conditions (Hm: males exposed to hypoxia; Hf: females exposed to hypoxia; Nm: males exposed to normoxia; Nf: females exposed to normoxia). Body weight at hatching and at 14-15 weeks was not affected by in ovo exposure to hypoxia. Mean arterial pressure and heart rate were not significantly altered by chronic in ovo hypoxia. However, isolated femoral arteries were more sensitive to electrical stimulation (frequency in Hz of half-maximal contraction, Hm: 1.62+/-0.33, Hf: 1.92+/-0.88, Nm: 2.49+/-0.49, Nf: 2.83+/-0.31) and pharmacological stimulation of peri-arterial sympathetic nerves (contraction in N/m in response to tyramine: Hm: 5.27+/-0.85, Hf: 4.10+/-0.9, Nm: 2.26+/-0.67, Nf: 3.65+/-0.51, p=0.07) after in ovo hypoxia. In side branches of the femoral artery, the effect of NO synthase blockade with L-NAME on contraction (in N/m) in response to high K+ (Hm: 0.35+/-0.91, Hf: 1.29+/-0.36, Nm: 2.88+/-0.19, Nf: 2.79+/-0.58) and on the sensitivity to acetylcholine (DeltapD2, H: 0.32+/-0.11, N: 0.62+/-0.05) was reduced after in ovo hypoxia. The present study shows that exposure to chronic moderate hypoxia during development affects the contractile and relaxing arterial responses of 14- to 15-week-old chickens. Although hypoxia did not lead to changes in blood pressure at this age, the observed effects on arterial sympathetic and endothelial function may represent early signs of future cardiovascular abnormalities.

Preferential renal and mesenteric vasodilation induced by barnidipine and amlodipine in spontaneously hypertensive rats.

Barnidipine is a stereoselective single isomer formulation of a long-term acting dihydropyridine calcium antagonist (CaA). In anaesthetised animals, the antihypertensive response to barnidipine is accompanied by a diuretic effect. The aim of the present study was to examine whether barnidipine increased renal blood flow in a conscious animal model for essential hypertension. We compared the regional specific hemodynamic effects of barnidipine with those obtained with its racemic mixture and amlodipine. Male adult spontaneously hypertensive rats (SHR) were instrumented with Doppler flow probes and catheters to measure renal (RVR), mesenteric (MVR) and hindquarter (HQVR) vascular resistance changes. One week after surgery, barnidipine, its racemic mixture, and amlodipine were intravenously administered at three doses (n> or =10 per dose) causing comparable reductions in mean arterial pressure (MAP). At doses of 3, 10 and 30 microg/kg barnidipine reduced MAP (+/- SEM) by 8+/-2, 26+/-3 and 45+/-4 mmHg. Equipotent effects on MAP were achieved by the racemic mixture of barnidipine at 10, 30 and 100 microg/kg, and by amlodipine at doses of 100, 300 and 1000 microg/kg. Following the 3 microg/kg and 10 microg/kg dose, barnidipine reduced MVR (% +/- SEM) by 4+/-4 and 19+/-4, and RVR by 8+/-2 and 15+/-4, respectively. In contrast, HQVR remained unaltered. Similar data were obtained for the racemic mixture of barnidipine and for amlodipine, although for the latter the changes in RVR were half of those found after barnidipine. After the highest doses of barnidipine, its racemic mixture as well as amlodipine, HQVR fell more than 25% whereas RVR and MVR remained unaltered. Analysis of the dynamic response to the CaAs revealed that the reductions in vascular resistance were associated with decreased myogenic-like oscillations in blood flow. We conclude that, in conscious SHR, the single isomer barnidipine reduces MAP at doses which are three times lower than its racemic mixture and 30 times lower than amlodipine. In contrast to short-acting CaAs such as nifedipine and isradipine, which reduce mainly HQVR and do not reduce RVR (Nievelstein et al.; Eur J Pharmacol 113:187-198, 1985), the three long-term acting CaAs preferentially dilated the mesenteric and renal vascular bed. In view of the elevation of RVR in essential hypertension, the reduction of RVR may contribute to the long-term antihypertensive effects of barnidipine and amlodipine.

17beta-estradiol attenuates the development of pressure-overload hypertrophy.

BACKGROUND: Cardiac hypertrophy is an independent risk factor for cardiovascular morbidity and mortality in men and in women. Epidemiological studies indicate that estrogen replacement therapy is cardioprotective; the mechanisms involved in this process, however, are poorly understood. We therefore studied the effect of 17beta-estradiol (E(2)) on the development of pressure-overload hypertrophy. METHODS AND RESULTS: Ovariectomized mice receiving E(2) or placebo underwent transverse aortic constriction (TAC) or sham operation. TAC led to a significant increase in ventricular mass compared with sham operation. E(2) treatment reduced cardiac hypertrophy by 31% and 26% compared with placebo 4 and 8 weeks after TAC, whereas it had no effect on the degree of pressure overload, as determined by hemodynamic measurements. Furthermore, E(2) blocked the increased phosphorylation of p38-mitogen-activated protein kinase (MAPK) observed in the placebo-treated animals with TAC. No differences were observed in the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK) 1/2 between the groups. E(2) had no effect on the expression of angiotensin-converting enzyme (ACE) or the angiotensin II type 1 receptor. Ventricular atrial natriuretic peptide (ANP) expression was detected only in the animals with TAC. Compared with placebo, E(2) treatment led to an increased expression of ANP in animals with pressure overload. CONCLUSIONS: Here, we show that E(2) attenuates the hypertrophic response to pressure overload in mice. This observation demonstrates that hormone replacement therapy with E(2) has direct effects on the heart and may be beneficial in the treatment of postmenopausal women to reduce cardiac hypertrophy.

Alterations in blood pressure and heart rate variability in transgenic rats with low brain angiotensinogen.

To study whether the brain renin-angiotensin system plays a role in the long-term and short-term control of blood pressure and heart rate variability, we examined in transgenic rats [TGR(ASrAOGEN)] with low brain angiotensinogen levels the 24-hour variation of blood pressure and heart rate. Telemetry recordings were made during basal and hypertensive conditions induced by a low-dose subcutaneous infusion of angiotensin II for 7 days. Short-term blood pressure and heart rate variability were evaluated by spectral analysis, and as a measure of baroreflex sensitivity, the average transfer gain between the pressure and heart rate variations was calculated. During the angiotensin II infusion in control but not TGR(ASrAOGEN) rats, the 24-hour rhythm of blood pressure was inverted (5.8+/-2 versus -0.4+/-1.8 mm Hg/group of day-night differences of blood pressure, P<0.05, respectively). In both the control and TGR(ASrAOGEN) rats, the 24-hour heart rate rhythms remained unaltered and paralleled those of locomotor activity. The transfer gain between 0.3 to 0.6 Hz was significantly higher in TGR(ASrAOGEN) than in control rats during control (0.71+/-0.1 versus 0.35+/-0.06, P<0.05) but not during angiotensin II infusion (0.6+/-0.07 versus 0.4+/-0.1, P>0.05). These results demonstrate that the brain renin-angiotensin system plays an important role in mediating the effects of angiotensin II on the circadian variation of blood pressure. Furthermore, these data indicate that a permanent deficiency in the brain renin-angiotensin system alters the reflex control of heart rate in rats.

Short-term and long-term blood pressure and heart rate variability in the mouse.

Knowledge on murine blood pressure and heart rate control mechanisms is limited. With the use of a tethering system, mean arterial pressure (MAP) and pulse interval (PI) were continuously recorded for periods up to 3 wk in Swiss mice. The day-to-day variation of MAP and PI was stable from 5 days after surgery. Within each mouse (n = 9), MAP and PI varied by 21+/-6 mm Hg and 17+/-4 ms around their respective 24-h averages (97+/-3 mm Hg and 89+/-3 ms). Over 24-h periods, MAP and PI were bimodally distributed and clustered around two preferential states. Short-term variability of MAP and PI was compared between the resting (control) and active states using spectral analysis. In resting conditions, variability of MAP was mainly confined to frequencies <1 Hz, whereas variability of PI was predominantly linked to the respiration cycle (3-6 Hz). In the active state, MAP power increased in the 0.08- to 3-Hz range, whereas PI power fell in the 0.08- to 0.4-Hz range. In both conditions, coherence between MAP and PI was high at 0.4 Hz with MAP leading the PI fluctuations by 0.3-0.4 s, suggesting that reflex coupling between MAP and PI occurred at the same frequency range as in rats. Short-term variability of MAP and PI was studied after intravenous injection of autonomic blockers. Compared with the resting control state, MAP fell and PI increased after ganglionic blockade with hexamethonium. Comparable responses of MAP were obtained with the alpha-blocker prazosin, whereas the beta-blocker metoprolol increased PI similarly. Muscarinic blockade with atropine did not significantly alter steady-state levels of MAP and PI. Both hexamethonium and prazosin decreased MAP variability in the 0.08- to 1-Hz range. In contrast, after hexamethonium and metoprolol, PI variability increased in the 0.4- to 3-Hz range. Atropine had no effect on MAP fluctuations but decreased those of PI in the 0.08- to 1-Hz range. These data indicate that, in mice, blood pressure and its variability are predominantly under sympathetic control, whereas both vagal and sympathetic nerves control PI variability. Blockade of endogenous nitric oxide formation by N(G)-nitro-L-arginine methyl ester increased MAP variability specifically in the 0.08- to 0.4-Hz range, suggesting a role of nitric oxide in buffering blood pressure fluctuations.

Timed inhibition of the renin-angiotensin system suppresses the rise in blood pressure upon awakening in spontaneously hypertensive rats.

In this study we investigated whether timed administration of drugs that inhibit the renin-angiotensin system can be used to blunt the rise in blood pressure that occurs during the transition from the resting to the active period of the day. For this purpose we compared in spontaneously hypertensive rats (SHR) the antihypertensive efficacy of the angiotensin converting enzyme (ACE) inhibitors captopril (doses: 3, 10, and 30 mg/ kg/6 h) and enalaprilat (0.3 mg/kg/6 h), and the AT1-receptor antagonist losartan (10 mg/kg/6 h) at two different treatment regimens. The antihypertensive drugs were given as a continuous 6-h infusion either during the transition from the dark to light period (DL) or that from the light to dark period (LD) for 5 consecutive days. For all agents, the average 24-h reduction of blood pressure was comparable for the LD or DL treatment regimen. However, the dynamics of the antihypertensive response were markedly different. The increase in blood pressure at awakening could be blunted much more effectively by the LD than DL treatment regimen. Furthermore, as indicated by the trough:peak ratios, blood pressure profiles were flatter with the LD than with the DL regimen. Thus, in SHR, 24-h rhythms of blood pressure can be modulated by timed administration of ACE inhibitors and losartan, such that the early morning rise in blood pressure is suppressed.

Phylogenetic relationships and evolution of the KNOTTED class of plant homeodomain proteins.

Knotted-like (KNOX) proteins constitute a group of homeodomain proteins involved in pattern formation in developing tissues of angiosperms and other green plants. We conducted phylogenetic analyses of nucleotide and amino acid sequences of all known KNOX proteins in order to examine their evolution. Our analyses reveal two groups of KNOX proteins, classes I and II. Dicot and monocot sequences occur in both classes, indicating that the protein classes arose prior to the origin of the monocots. A conifer (Picea) sequence is nested within class I, suggesting that there are likely to be other copies of KNOX genes in this and other conifers. The orthology of several grass genes (including Zea Kn1, ZMKN1) is strongly supported by phylogenetic and synteny analyses. However, no compelling evidence supports the hypothesis of orthology previously proposed for several dicot genes and ZMKN1. Analysis of expression patterns suggests that the ancestral KNOX gene was expressed in all plant parts and that the propensity to be downregulated in roots and leaves evolved in the class I genes.

Isolation and characterization of two knotted-like homeobox genes from tomato.

Homeobox genes are known to play a role in developmental regulation. The knotted-like homeobox (knox) genes fall into two classes. The class I knox genes like knl, stml, and knatl are involved in maintaining meristem identity in cells. The function of class II knox genes is at yet undetermined. We have characterized two knox genes from tomato. LeT6 and LeT12 map to distinct chromosome locations that are different from the location for a recently cloned knox gene from tomato, tknl, confirming that plant homeobox genes are not clustered on chromosomes. These genes have a distinct expression pattern. Unlike other class I knl-like genes, LeT6 is expressed in developing lateral organs and developing ovaries in flowers. LeT12 is more ubiquitously expressed in the mature plant. RNA in situ localization data suggest that both these genes may have a role to play in formative events in ovule and embryo morphogenesis.

Blood pressure and heart rate variability in early pregnancy in rats.

Changes in the autonomic control of the circulation may contribute to the maternal hemodynamic adaptation to early pregnancy. To evaluate this, we studied mean arterial pressure (MAP) and heart rate (HR) in chronically instrumented, conscious rats in early (days 4, 6, 8, and 10) and late (day 18) pregnancy (n = 8) and in nonpregnant rats (n = 9). MAP and HR were recorded on a beat-to-beat basis and analyzed by spectral analysis. Spectral density power was calculated in low- (0.047-0.305 Hz), mid- (0.305-0.598 Hz), and high-frequency (0.598-1.494 Hz) bands, which contain oscillations that are among others related to myogenic-, sympathetic/vagal-, and vagal/respiration-related influences, respectively. In addition, baroreceptor reflex sensitivity was determined from spontaneous variations in MAP and HR by a sequential time series method and by calculating the transfer gain between MAP and HR in the midfrequency band. Mean values of HR and MAP did not differ between the two groups on day 4. In the pregnant group, MAP fell gradually over days, whereas HR had significantly increased only on day 18. Overall variability in MAP and HR (expressed as coefficients of variation) did not change during pregnancy. Baroreceptor reflex sensitivity did not differ between the groups and did not change with advancing pregnancy. Spontaneous oscillations of MAP and HR at low, mid, and high frequencies were not different between pregnant and nonpregnant rats on days 4 to 10. On day 18, spectral density power of MAP, but not of HR, in the high-frequency band had significantly increased in pregnant rats only, most likely reflecting the increased impact of breathing on MAP fluctuations. We conclude that, with the methods employed, we could not discern any changes in baroreflex sensitivity and MAP and HR variability in pregnancy. This would imply that changes in autonomic activity do not contribute appreciably to the hemodynamic adaptations in early rat pregnancy.

A gene fusion at a homeobox locus: alterations in leaf shape and implications for morphological evolution.

Compound leaves are seen in many angiosperm genera and are thought to be either fundamentally different from simple leaves or elaborations of simple leaves. The knotted1-like homeobox (knox) genes are known to regulate plant development. When overexpressed in homologous or heterologous species, this family of genes can cause changes in leaf morphology, including excessive leaf compounding in tomato. We describe here an instance of a spontaneously arisen fusion between a gene encoding a metabolic enzyme and a homeodomain protein. We show that the fusion results in overexpression of the homeodomain protein and a change in morphology that approximates the changes caused by overexpression of the same gene under the control of the cauliflower mosaic virus 35S promoter in transgenic plants. Exon-shuffling events can account for the modularity of proteins. If the shuffled exons are associated with altered promoters, changes in gene expression patterns can result. Our results show that gene fusions of this nature can cause changes in expression patterns that lead to altered morphology. We suggest that such phenomena may have played a role in the evolution of form.

Frequency-dependent modulation of renal blood flow by renal nerve activity in conscious rabbits.

To examine the influence of the various frequency components of renal sympathetic nerve activity (RSNA) on renal blood flow (RBF) dynamics, a Doppler flow probe and renal nerve electrode were implanted on the left renal artery of 10 rabbits. Experiments were performed 4-9 days after surgery. Physiological changes in RSNA were induced by subjecting the rabbits to periods of breathing hypoxic gas mixtures. Signals were sampled at 1 kHz and analyzed by spectral analysis. During moderate hypoxia (arterial PO2 = 44 +/- 1 mmHg), arterial pressure and heart rate did not change, averaged RSNA increased by 90 +/- 7%, and RBF fell by 18 +/- 3%. In a separate group of renal-denervated rabbits (n = 6), no changes in RBF occurred during hypoxia. In intact rabbits, 53 +/- 4% of spectral density power of RSNA was found at the cardiac frequency and the remainder was predominantly coupled to respiration (approximately 0.9 Hz). During moderate hypoxia the amplitude of the RSNA oscillations increased 17 +/- 6 times at the cardiac frequency and 10 +/- 3 times at the respiration-related frequency. Modulation of RBF variability by the fluctuations of RSNA at the cardiac- and respiration-related frequency was, however, small. The normalized transfer gain between RSNA and RBF was approximately 0.1 at > 0.5 Hz. This means that, at > 0.5 Hz, maximally 10% of the amplitude of the RSNA oscillations is transmitted to corresponding RBF fluctuations. These transfer properties did not change during hypoxia. At < 0.5 Hz the transfer gain between RSNA and RBF increased. During moderate hypoxia, 0.3-Hz coherent oscillations of RSNA and RBF were found. In renal-denervated rabbits, 0.3-Hz oscillations in RBF were absent. Thus the renal vasculature was able to follow relatively low-frequency (< 0.5-Hz) fluctuations of RSNA and responded with corresponding oscillations in RBF. In contrast, the renal vasculature responded with increased constriction at the high-frequency (> 0.5-Hz) fluctuations of RSNA. These findings suggest that, in conscious rabbits, high-frequency oscillations of RSNA contribute to the vasoconstrictor tone, whereas the lower frequencies of RSNA contribute to the variability of RBF.