The longevity gene mIndy (I'm Not Dead, Yet) affects blood pressure through sympathoadrenal mechanisms.

Reduced expression of the plasma membrane citrate transporter INDY (acronym I'm Not Dead, Yet) extends life span in lower organisms. Deletion of the mammalian Indy (mIndy) gene in rodents improves metabolism via mechanisms akin to caloric restriction, known to lower blood pressure (BP) by sympathoadrenal inhibition. We hypothesized that mIndy deletion attenuates sympathoadrenal support of BP. Continuous arterial BP and heart rate (HR) were reduced in mINDY-KO mice. Concomitantly, urinary catecholamine content was lower, and the decreases in BP and HR by mIndy deletion were attenuated after autonomic ganglionic blockade. Catecholamine biosynthesis pathways were reduced in mINDY-KO adrenals using unbiased microarray analysis. Citrate, the main mINDY substrate, increased catecholamine content in pheochromocytoma cells, while pharmacological inhibition of citrate uptake blunted the effect. Our data suggest that deletion of mIndy reduces sympathoadrenal support of BP and HR by attenuating catecholamine biosynthesis. Deletion of mIndy recapitulates beneficial cardiovascular and metabolic responses to caloric restriction, making it an attractive therapeutic target.

C-type natriuretic peptide and natriuretic peptide receptor B signalling inhibits cardiac sympathetic neurotransmission and autonomic function.

AIMS: B-type natriuretic peptide (BNP)-natriuretic peptide receptor A (NPR-A) receptor signalling inhibits cardiac sympathetic neurotransmission, although C-type natriuretic peptide (CNP) is the predominant neuropeptide of the nervous system with expression in the heart and vasculature. We hypothesized that CNP acts similarly to BNP, and that transgenic rats (TGRs) with neuron-specific overexpression of a dominant negative NPR-B receptor would develop heightened sympathetic drive. METHODS AND RESULTS: Mean arterial pressure and heart rate (HR) were significantly (P < 0.05) elevated in freely moving TGRs (n = 9) compared with Sprague Dawley (SD) controls (n = 10). TGR had impaired left ventricular systolic function and spectral analysis of HR variability suggested a shift towards sympathoexcitation. Immunohistochemistry demonstrated co-staining of NPR-B with tyrosine hydroxylase in stellate ganglia neurons. In SD rats, CNP (250 nM, n = 8) significantly reduced the tachycardia during right stellate ganglion stimulation (1-7 Hz) in vitro whereas the response to bath-applied norepinephrine (NE, 1 µM, n = 6) remained intact. CNP (250 nM, n = 8) significantly reduced the release of 3H-NE in isolated atria and this was prevented by the NPR-B antagonist P19 (250 nM, n = 6). The neuronal Ca2+ current (n = 6) and intracellular Ca2+ transient (n = 9, using fura-2AM) were also reduced by CNP in isolated stellate neurons. Treatment of the TGR (n = 9) with the sympatholytic clonidine (125 µg/kg per day) significantly reduced mean arterial pressure and HR to levels observed in the SD (n = 9). CONCLUSION: C-type natriuretic peptide reduces cardiac sympathetic neurotransmission via a reduction in neuronal calcium signalling and NE release through the NPR-B receptor. Situations impairing CNP-NPR-B signalling lead to hypertension, tachycardia, and impaired left ventricular systolic function secondary to sympatho-excitation.

Cholinergic Pathway Suppresses Pulmonary Innate Immunity Facilitating Pneumonia After Stroke.

BACKGROUND AND PURPOSE: Temporary immunosuppression has been identified as a major risk factor for the development of pneumonia after acute central nervous system injury. Although overactivation of the sympathetic nervous system was previously shown to mediate suppression of systemic cellular immune responses after stroke, the role of the parasympathetic cholinergic anti-inflammatory pathway in the antibacterial defense in lung remains largely elusive. METHODS: The middle cerebral artery occlusion model in mice was used to examine the influence of the parasympathetic nervous system on poststroke immunosuppression. We used heart rate variability measurement by telemetry, vagotomy, α7 nicotinic acetylcholine receptor-deficient mice, and parasympathomimetics (nicotine, PNU282987) to measure and modulate parasympathetic activity. RESULTS: Here, we demonstrate a rapidly increased parasympathetic activity in mice after experimental stroke. Inhibition of cholinergic signaling by either vagotomy or by using α7 nicotinic acetylcholine receptor-deficient mice reversed pulmonary immune hyporesponsiveness and prevented pneumonia after stroke. In vivo and ex vivo studies on the role of α7 nicotinic acetylcholine receptor on different lung cells using bone marrow chimeric mice and isolated primary cells indicated that not only macrophages but also alveolar epithelial cells are a major cellular target of cholinergic anti-inflammatory signaling in the lung. CONCLUSIONS: Thus, cholinergic pathways play a pivotal role in the development of pulmonary infections after acute central nervous system injury.

Preserved functional autonomic phenotype in adult mice overexpressing moderate levels of human alpha-synuclein in oligodendrocytes.

Mice overexpressing human alpha-synuclein in oligodendrocytes (MBP1-α-syn) recapitulate some key functional and neuropathological features of multiple system atrophy (MSA). Whether or not these mice develop severe autonomic failure, which is a key feature of human MSA, remains unknown. We explored cardiovascular autonomic regulation using long-term blood pressure (BP) radiotelemetry and pharmacological testing. We instrumented 12 MBP1-α-syn mice and 11 wild-type mice aged 9 months for radiotelemetry. Animals were tested with atropine, metoprolol, clonidine, and trimethaphan at 9 and 12 months age. We applied spectral and cross-spectral analysis to assess heart rate (HR) and BP variability. At 9 months of age daytime BP (transgenic: 101 ± 2 vs. wild type: 99 ± 2 mmHg) and HR (497 ± 11 vs. 505 ± 16 beats/min) were similar. Circadian BP and HR rhythms were maintained. Nighttime BP (109 ± 2 vs. 108 ± 2 mmHg) and HR (575 ± 15 vs. 569 ± 14 beats/min), mean arterial BP responses to trimethaphan (-21 ± 8 vs. -10 ± 5 mmHg, P = 0.240) and to clonidine (-8 ± 3 vs. -5 ± 2 mmHg, P = 0.314) were similar. HR responses to atropine (+159 ± 24 vs. +146 ± 22 beats/min), and to clonidine (-188 ± 21 vs. -163 ± 33 beats/min) did not differ between strains. Baroreflex sensitivity (4 ± 1 vs. 4 ± 1 msec/mmHg) and HR variability (total power, 84 ± 17 vs. 65 ± 21 msec²) were similar under resting conditions and during pharmacological testing. Repeated measurements at 12 months of age provided similar results. In mice, moderate overexpression of human alpha-synuclein in oligodendrocytes is not sufficient to induce overt autonomic failure. Additional mechanisms may be required to express the autonomic failure phenotype including higher levels of expression or more advanced age.

AVE 0991, a non-peptide Mas-receptor agonist, facilitates penile erection.

The renin-angiotensin system plays a crucial role in erectile function. It has been shown that elevated levels of angiotensin II contribute to the development of erectile dysfunction both in humans and in aminals. On the contrary, the heptapeptide angiotensin-(1-7) appears to mediate penile erection by activation of the Mas receptor. Recently, we have shown that the erectile function of Mas gene-deleted mice was substantially reduced, which was associated with a marked increase in fibrous tissue in the corpus cavernosum. We have hypothesized that the synthetic non-peptide Mas agonist, AVE 0991, would potentiate penile erectile function. We showed that intracavernosal injection of AVE 0991 potentiated the erectile response of anaesthetized Wistar rats, measured as the ratio between corpus cavernosum pressure and mean arterial pressure, upon electrical stimulation of the major pelvic ganglion. The facilitatory effect of AVE 0991 on erectile function was dose dependent and completely blunted by the nitric oxide synthesis inhibitor, l-NAME. Importantly, concomitant intracavernosal infusion of the specific Mas receptor blocker, A-779, abolished the effect of AVE 0991. We demonstrated that AVE 0991 potentiates the penile erectile response through Mas in an NO-dependent manner. Importantly, these results suggest that Mas agonists, such as AVE 0991, might have significant therapeutic benefits for the treatment of erectile dysfunction.

Spinophilin regulates central angiotensin II-mediated effect on blood pressure.

Central angiotensin II (AngII) plays an important role in the regulation of the sympathetic nervous system. The underlining molecular mechanisms are largely unknown. Spinophilin (SPL) is a regulator of G protein-coupled receptor signaling. Deletion of SPL induces sympathetically mediated arterial hypertension in mice. We tested the hypothesis that SPL restrains blood pressure (BP) by regulating AngII activity. We equipped SPL(-/-) and SPL(+/+) mice with telemetric devices and applied AngII (1.0 mg kg(-1) day(-1), minipumps) or the AngII subtype 1 receptor (AT1-R) blocker valsartan (50 mg kg(-1) day(-1), gavage). We assessed autonomic nervous system activity through intraperitoneal application of trimethaphan, metoprolol, and atropine. We also tested the effect of intracerebroventricular (icv) AngII on blood pressure in SPL(-/-) and in SPL(+/+) mice. Chronic infusion of AngII upregulates SPL expression in the hypothalamus of SPL(+/+) mice. Compared with SPL(+/+) mice, SPL(-/-) mice showed a greater increase in daytime BP with AngII (19.2 ± 0.8 vs. 13.5 ± 1.6 mmHg, p < 0.05). SPL(-/-) showed a greater depressor response to valsartan. BP and heart rate decreased more with trimethaphan and metoprolol in AngII-treated SPL(-/-) than in AngII-treated SPL(+/+) mice. SPL(-/-) mice responded more to icv AngII. Furthermore, brainstem AT1-R and AngII type 2 receptor (AT2-R) expression was reduced in SPL(-/-) mice. AngII treatment normalized AT1-R and AT2-R expression levels. In summary, our findings suggest that SPL restrains AngII-mediated sympathetic nervous system activation. SPL is a hitherto unrecognized molecule with regard to central blood pressure control and may pave the way to novel strategies for the treatment of hypertension.

Estrogen receptor-beta signals left ventricular hypertrophy sex differences in normotensive deoxycorticosterone acetate-salt mice.

We found earlier that deoxycorticosterone acetate-salt treatment causes blood pressure-independent left ventricular hypertrophy, but only in male mice. To test the hypothesis that the estrogen receptor-ß (ERß) protects the females from left ventricular hypertrophy, we treated male and female ERß-deficient (ERß(-/-)) mice and their male and female littermates (wild-type [WT]) with deoxycorticosterone acetate-salt and made them telemetrically normotensive with hydralazine. WT males had increased (+16%) heart weight/tibia length ratios compared with WT females (+7%) at 6 weeks. In ERß(-/-) mice, this situation was reversed. Female WT mice had the greatest heart weight/tibia length ratio increases of all of the groups (+23%), even greater than ERß(-/-) males (+10%). Echocardiography revealed concentric left ventricular hypertrophy in male WT mice, whereas ERß(-/-) females developed dilative left ventricular hypertrophy. The hypertrophic response in female ERß(-/-) mice was accompanied by the highest degree of collagen deposition, indicating maladaptive remodeling. ERß(+/+) females showed robust protective p38 and extracellular signal-regulated kinase 1/2 signaling relationships compared with other groups. Calcineurin Aß expression and its positive regulator myocyte-enriched calcineurin-interacting protein 1 were increased in deoxycorticosterone acetate-salt female ERß(-/-) mice, yet lower than in WT males. Endothelin increased murine cardiomyocyte hypertrophy in vitro, which could be blocked by estradiol and an ERß agonist. We conclude that a functional ERß is essential for inducing adaptive p38 and extracellular signal-regulated kinase signaling, while reducing maladaptive calcineurin signaling in normotensive deoxycorticosterone acetate female mice. Our findings address the possibility of sex-specific cardiovascular therapies.

Role of the multidomain protein spinophilin in blood pressure and cardiac function regulation.

Spinophilin controls intensity/duration of G protein-coupled receptor signaling and thereby influences synaptic activity. We hypothesize that spinophilin affects blood pressure through central mechanisms. We measured blood pressure and heart rate in SPL-deficient (SPL(-/-)), heterozygous SPL-deficient (SPL(+/-)), and wild-type (SPL(+/+)) mice by telemetry combined with fast Fourier transformation. We also assessed peripheral vascular reactivity and performed echocardiography. SPL(-/-) had higher mean arterial pressure than SPL(+/-) and SPL(+/+) (121+/-2, 112+/-1, and 113+/-1 mm Hg). Heart rate was inversely related to spinophilin expression (SPL(-/-) 565+/-0.4, SPL(+/-) 541+/-5, SPL(+/+) 525+/-8 bpm). The blood pressure response to prazosin, trimethapane, and the heart rate response to metoprolol were stronger in SPL(-/-) than SPL(+/+) mice, whereas heart rate response to atropine was attenuated in SPL(-/-). Mesenteric artery vasoreactivity after angiotensin II, phenylephrine, and the thromboxane mimetic (U46619) as well as change in heart rate, stroke volume, and cardiac output after dobutamine were similar in SPL(-/-) and SPL(+/+). Baroreflex sensitivity was attenuated in SPL(-/-) compared with SPL(+/-) and SPL(+/+), which was confirmed by pharmacological testing. Heart rate variability parameters were attenuated in SPL(-/-) mice. We suggest that an increase in central sympathetic outflow participates in blood pressure and heart rate increases in SPL(-/-) mice. The elevated blood pressure in SPL(-/-) mice was associated with attenuated baroreflex sensitivity and decreased parasympathetic activity. Our study is the first to show a role for the spinophilin gene in blood pressure regulation.

Deoxycorticosterone acetate-salt mice exhibit blood pressure-independent sexual dimorphism.

We tested the hypothesis that female and male mice differ in terms of cardiac hypertrophy after deoxycorticosterone acetate (DOCA)+salt hypertension (uninephrectomy and 1% saline in drinking water) and focused on calcineurin signaling. We excluded confounding effects of blood pressure elevation or sex-related blood pressure differences by treating DOCA-salt mice with hydralazine (250 mg/L in drinking water). We found that directly measured mean arterial blood pressure was lowered to control values with hydralazine and corroborated this finding in separate mouse groups with radiotelemetry. Male mice were more responsive to DOCA-salt-related effects. They developed more left ventricular hypertrophy and more renal hypertrophy after 6 weeks of DOCA-salt+hydralazine compared with female mice. In hearts, transcripts for calcineurin Abeta and for myocyte-enriched calcineurin interacting protein 1 were upregulated in male but not in female mice. Enhanced activity of calcineurin Abeta, as indicated by diminished phosphorylation of NFATc2 in male mice, accounted for this sex-specific difference. Stretch-related, inflammatory, and profibrotic responses were also accentuated in male mice, as shown by higher transcript levels of atrial natriuretic peptide, monocyte chemoattractant protein-1, and transforming growth factor-beta. Our results support sex-specific regulation of the calcineurin pathway in response to largely blood pressure-independent mineralocorticoid action. We suggest that sex-specific calcineurin activation determines the maladaptive cardiac and renal hypertrophic responses and accompanying organ injury in male mice.

Cardiovascular autonomic regulation in Non-Obese Diabetic (NOD) mice.

Non-Obese Diabetic (NOD) mice show profound pathomorphological changes in sympathetic ganglia during the development of type 1 diabetes mellitus. We tested the hypothesis that NOD mice represent an experimental model to investigate cardiovascular changes seen in humans with diabetic autonomic neuropathy. Blood glucose (BG) levels were measured once a week. Diabetes mellitus was diagnosed as BG levels exceeded 250 mg/dl twice. NOD mice that did not become diabetic served as control group. Blood pressure (BP) and heart rate (HR) were monitored by telemetry and baroreflex sensitivity (BRS) was calculated with the sequence method or with cross spectral analysis. The measurements were obtained before onset of diabetes and during the 4th week of diabetes. The onset of diabetes was accompanied by a continuous decline in HR (615+/-14 vs. 498+/-23 bpm), whereas BP values remained stable (108+/-2 vs. 111+/-2 mm Hg). The circadian HR rhythm increased in diabetic NOD mice. BRS was higher in diabetic NOD mice than in controls. Atropine reduced BRS more profoundly in diabetic mice compared to non-diabetic mice. Despite pathomorphological similarities of the diabetic autonomic neuropathy between patients with diabetes and diabetic NOD mice, the changes in blood pressure regulation are different. In conclusion the use of diabetic NOD mice as a functional model for human diabetes may be questioned.

Evidence that the vasodilator angiotensin-(1-7)-Mas axis plays an important role in erectile function.

The vasodilator/antiproliferative peptide angiotensin-(1-7) [ANG-(1-7)] is released into the corpus cavernosum sinuses, but its role in erectile function has yet to be defined. In this study, we sought to determine whether ANG-(1-7) and its receptor Mas play a role in erectile function. The ANG-(1-7) receptor Mas was immunolocalized in rat corpus cavernosum by confocal microscopy. Infusion of ANG-(1-7) into corpus cavernosum at a rate of 15.5 pmol x kg(-1) x min(-1) potentiated the elevation of the corpus cavernosum pressure induced by electrical stimulation of the major pelvic ganglion (MPG) in rats. The facilitatory effect of ANG-(1-7) was completely blunted by the specific ANG-(1-7) receptor blocker A-779 and N(omega)-nitro-L-arginine methyl ester. Nitric oxide (NO) release in the corpus cavernosum was evaluated with the fluorescent dye 4-amino-5 methylamino-2',7'-difluorofluorescein diacetate. Electrical stimulated-release of NO in rat corpus cavernosum was potentiated by ANG-(1-7). Furthermore, incubation of rat and mouse corpus cavernosum strips with ANG-(1-7) at 10 nmol/l resulted in an increase of NO release. This effect was completely abolished in mas-deficient mice. More importantly, genetic deletion of Mas resulted in compromised erectile function as demonstrated by penile fibrosis and severely depressed response to electrical stimulation of the MPG. Furthermore, the attenuated erectile function of DOCA-salt hypertensive rats was fully restored by ANG-(1-7) administration. Together these data provide strong evidence for a key role of the ANG-(1-7)-Mas axis in erectile function.

Regulator of G protein signalling 2 ameliorates angiotensin II-induced hypertension in mice.

Angiotensin II (Ang II) activates signalling pathways predominantly through the G-protein-coupled Ang II type 1 receptor (AT(1)R). The regulator of G protein signalling 2 (RGS2) is a negative G protein regulator. We hypothesized that RGS2 deletion changes blood pressure regulation by increasing the response to Ang II. To address this issue, we infused Ang II (0.5 mg kg(-1) day(-1)) chronically into conscious RGS2-deleted (RGS2(-/-)) and wild-type (RGS2(+/+)) mice, measured mean arterial blood pressure and heart rate (HR) with telemetry and assessed vasoreactivity and gene expression of AT(1A), AT(1B) and AT(2) receptors. Angiotensin II infusion increased blood pressure more in RGS2(-/-) than in RGS2(+/+) mice, while HR was not different between the groups, indicating a resetting of the baroreceptor reflex. Urinary catecholamine excretion was similar in Ang II-infused RGS2(-/-) and RGS2(+/+) mice, indicating a minor role of sympathetic tone for blood pressure differences. Myogenic tone and vasoreactivity in response to Ang II, endothelin-1 and phenylephrine were increased in isolated renal interlobar arterioles of RGS2(-/-) mice compared with RGS2(+/+) mice. The AT(1A), AT(1B) and AT(2) receptor gene expression was not different between RGS2(-/-) and RGS2(+/+) mice. Our findings suggest that RGS2 deletion promotes Ang II-dependent hypertension primarily through an increase of myogenic tone and vasoreactivity, probably by sensitization of AT(1) receptors.