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1.
Circ Heart Fail ; 14(12): e008365, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34789005

RESUMO

BACKGROUND: Recent clinical studies demonstrate that SGLT2 (sodium-glucose cotransporter 2) inhibitors ameliorate heart failure (HF). The present study was conducted to assess the expression and function of renal SGLT2 and the influence of enhanced renal sympathetic tone in HF. METHODS: Four weeks after coronary artery ligation surgery to induce HF, surgical bilateral renal denervation (RDN) was performed in rats. Four groups of rats (Sham-operated control [Sham], Sham+RDN, HF and HF+RDN; n=6/group) were used. Immunohistochemistry and Western blot analysis were performed to evaluate the renal SGLT2 expression. One week after RDN (5 weeks after induction of HF), intravenous injection of SGLT2 inhibitor dapagliflozin were performed to assess renal excretory responses. In vitro, human embryonic kidney cells were used to investigate the fractionation of SGLT2 after norepinephrine treatment. RESULTS: In rats with HF, (1) SGLT2 expression in the proximal tubule of the kidney was increased; (2) the response of increases in urine flow, sodium excretion, and glucose excretion to dapagliflozin were greater; and (3) RDN attenuated renal SGLT2 expression and normalized renal functional responses to dapagliflozin. In vitro, norepinephrine promoted translocation of SGLT2 to the cell membrane. CONCLUSIONS: These results indicate that the enhanced tonic renal sympathetic nerve activation in HF increases the expression and functional activity of renal SGLT2. Potentiated trafficking of SGLT2 to cell surface in renal proximal tubules mediated by norepinephrine may contribute to this functional activation of SGLT2 in HF. These findings provide critical insight into the underlying mechanisms for the beneficial effects of SGLT2 inhibitors on HF reported in the clinical studies.


Assuntos
Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Rim/inervação , Transportador 2 de Glucose-Sódio/metabolismo , Animais , Glucose/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Masculino , Ratos Sprague-Dawley , Sódio/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/fisiopatologia
2.
Hypertension ; 77(1): 147-157, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33296248

RESUMO

Central infusion of Ang II (angiotensin II) has been associated with increased sympathetic outflow resulting in neurogenic hypertension. In the present study, we appraised whether the chronic increase in central Ang II activates the paraventricular nucleus of the hypothalamus (PVN) resulting in elevated sympathetic tone and altered baro- and chemoreflexes. Further, we evaluated the contribution of HIF-1α (hypoxia-inducible factor-1α), a transcription factor involved in enhancing the expression of N-methyl-D-aspartate receptors and thus glutamatergic-mediated sympathetic tone from the PVN. Ang II infusion (20 ng/minute, intracerebroventricular, 14 days) increased mean arterial pressure (126±9 versus 84±4 mm Hg), cardiac sympathetic tone (96±7 versus 75±6 bpm), and decreased cardiac parasympathetic tone (16±2 versus 36±3 versus bpm) compared with saline-infused controls in conscious rats. The Ang II-infused group also showed an impaired baroreflex control of heart rate (-1.50±0.1 versus -2.50±0.3 bpm/mm Hg), potentiation of the chemoreflex pressor response (53±7 versus 30±7 mm Hg) and increased number of FosB-labeled cells (53±3 versus 19±4) in the PVN. Concomitant with the activation of the PVN, there was an increased expression of HIF-1α and N-Methyl-D-Aspartate-type1 receptors in the PVN. Further, Ang II-infusion showed increased renal sympathetic nerve activity (20.5±2.3% versus 6.4±1.9% of Max) and 3-fold enhanced renal sympathetic nerve activity responses to microinjection of N-methyl-D-aspartate (200 pmol) into the PVN of anesthetized rats. Further, silencing of HIF-1α in NG108 cells abrogated the expression of N-methyl-D-aspartate-N-methyl-D-aspartate-type1 induced by Ang II. Taken together, our studies suggest a novel Ang II-HIF-1α-N-methyl-D-aspartate receptor-mediated activation of preautonomic neurons in the PVN, resulting in increased sympathetic outflow and alterations in baro- and chemoreflexes.


Assuntos
Angiotensina II/farmacologia , Ácido Glutâmico/fisiologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/fisiologia , Núcleo Hipotalâmico Paraventricular/efeitos dos fármacos , Sistema Nervoso Simpático/efeitos dos fármacos , Animais , Barorreflexo/efeitos dos fármacos , Barorreflexo/fisiologia , Hipertensão/fisiopatologia , Rim/inervação , Masculino , N-Metilaspartato/farmacologia , Núcleo Hipotalâmico Paraventricular/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/fisiologia , Sistema Nervoso Simpático/fisiologia
3.
Am J Physiol Heart Circ Physiol ; 319(6): H1414-H1437, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33064567

RESUMO

Increased matrix metalloprotease 9 (MMP9) after myocardial infarction (MI) exacerbates ischemia-induced chronic heart failure (CHF). Autophagy is cardioprotective during CHF; however, whether increased MMP9 suppresses autophagic activity in CHF is unknown. This study aimed to determine whether increased MMP9 suppressed autophagic flux and MMP9 inhibition increased autophagic flux in the heart of rats with post-MI CHF. Sprague-Dawley rats underwent either sham surgery or coronary artery ligation 6-8 wk before being treated with MMP9 inhibitor for 7 days, followed by cardiac autophagic flux measurement with lysosomal inhibitor bafilomycin A1. Furthermore, autophagic flux was measured in vitro by treating H9c2 cardiomyocytes with two independent pharmacological MMP9 inhibitors, salvianolic acid B (SalB) and MMP9 inhibitor-I, and CRISPR/cas9-mediated MMP9 genetic ablation. CHF rats showed cardiac infarct, significantly increased left ventricular end-diastolic pressure (LVEDP), and increased MMP9 activity and fibrosis in the peri-infarct areas of left ventricular myocardium. Measurement of the autophagic markers LC3B-II and p62 with lysosomal inhibition showed decreased autophagic flux in the peri-infarct myocardium. Treatment with SalB for 7 days in CHF rats decreased MMP9 activity and cardiac fibrosis but increased autophagic flux in the peri-infarct myocardium. As an in vitro corollary study, measurement of autophagic flux in H9c2 cardiomyocytes and fibroblasts showed that pharmacological inhibition or genetic ablation of MMP9 upregulates autophagic flux. These data are consistent with our observations that MMP9 inhibition upregulates autophagic flux in the heart of rats with CHF. In conclusion, the results in this study suggest that the beneficial outcome of MMP9 inhibition in pathological cardiac remodeling is in part mediated by improved autophagic flux.NEW & NOTEWORTHY This study elucidates that the improved cardiac extracellular matrix (ECM) remodeling and cardioprotective effect of matrix metalloprotease 9 (MMP9) inhibition in chronic heart failure (CHF) are via increased autophagic flux. Autophagy is cardioprotective; however, the mechanism of autophagy suppression in CHF is unknown. We for the first time demonstrated here that increased MMP9 suppressed cardiac autophagy and ablation of MMP9 increased cardiac autophagic flux in CHF rats. Restoring the physiological level of autophagy in the failing heart is a challenge, and our study addressed this challenge. The novelty and highlights of this report are as follows: 1) MMP9 regulates cardiomyocyte and fibroblast autophagy, 2) MMP9 inhibition protects CHF after myocardial infarction (MI) via increased cardiac autophagic flux, 3) MMP9 inhibition increased cardiac autophagy via activation of AMP-activated protein kinase (AMPK)α, Beclin-1, Atg7 pathway and suppressed mechanistic target of rapamycin (mTOR) pathway.


Assuntos
Autofagia/efeitos dos fármacos , Benzofuranos/farmacologia , Fibroblastos/efeitos dos fármacos , Insuficiência Cardíaca/tratamento farmacológico , Metaloproteinase 9 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Animais , Linhagem Celular , Modelos Animais de Doenças , Fibroblastos/enzimologia , Fibroblastos/patologia , Fibrose , Insuficiência Cardíaca/enzimologia , Insuficiência Cardíaca/patologia , Insuficiência Cardíaca/fisiopatologia , Masculino , Metaloproteinase 9 da Matriz/genética , Camundongos , Miócitos Cardíacos/enzimologia , Miócitos Cardíacos/patologia , Ratos Sprague-Dawley , Transdução de Sinais , Função Ventricular Esquerda/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacos
4.
Neuropeptides ; 83: 102076, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32800589

RESUMO

The basolateral amygdala (BLA) is critical in the control of the sympathetic output during stress. Studies demonstrated the involvement of the renin-angiotensin system components in the BLA. Angiotensin-(1-7) [Ang-(1-7)], acting through Mas receptors, reduces stress effects. Considering that angiotensin-converting enzyme 2 (ACE2) is the principal enzyme for the production of Ang-(1-7), here we evaluate the cardiovascular reactivity to acute stress after administration of the ACE2 activator, diminazene aceturate (DIZE) into the BLA. We also tested whether systemic treatment with DIZE could modify synaptic activity in the BLA and its effect directly on the expression of the N-methyl-d-aspartate receptors (NMDARs) in NG108 neurons in-vitro. Administration of DIZE into the BLA (200 pmol/100 nL) attenuated the tachycardia to stress (ΔHR, bpm: vehicle = 103 ± 17 vs DIZE = 49 ± 7 p = 0.018); this effect was inhibited by Ang-(1-7) antagonist, A-779 (ΔHR, bpm: DIZE = 49 ± 7 vs A-779 + DIZE = 100 ± 15 p = 0.04). Systemic treatment with DIZE attenuated the excitatory synaptic activity in the BLA (Frequency (Hz): vehicle = 2.9 ± 0.4 vs. DIZE =1.8 ± 0.3 p < 0.04). NG108 cells treated with DIZE demonstrated decreased expression of l subunit NMDAR-NR1 (NR1 expression (a.u): control = 0.534 ± 0.0593 vs. DIZE = 0.254 ± 0.0260) of NMDAR and increases of Mas receptors expression. These data demonstrate that DIZE attenuates the tachycardia evoked by acute stress. This effect results from a central action in the BLA involving activation of Mas receptors. The ACE2 activation via DIZE treatment attenuated the frequency of excitatory synaptic activity in the basolateral amygdala and this effect can be related with the decreases of the NMDAR-NR1 receptor expression.


Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , Complexo Nuclear Basolateral da Amígdala/efeitos dos fármacos , Diminazena/análogos & derivados , Ácido Glutâmico/metabolismo , Frequência Cardíaca/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Taquicardia/metabolismo , Angiotensina I/antagonistas & inibidores , Angiotensina II/análogos & derivados , Angiotensina II/farmacologia , Animais , Complexo Nuclear Basolateral da Amígdala/metabolismo , Diminazena/farmacologia , Neurônios/metabolismo , Fragmentos de Peptídeos/antagonistas & inibidores , Fragmentos de Peptídeos/farmacologia , Ratos , Ratos Wistar , Receptores de N-Metil-D-Aspartato/metabolismo
6.
Cardiovasc Diabetol ; 19(1): 57, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32384887

RESUMO

BACKGROUND: Glucagon-like peptide-1 (GLP-1) induces diuresis and natriuresis. Previously we have shown that GLP-1 activates afferent renal nerve to increase efferent renal sympathetic nerve activity that negates the diuresis and natriuresis as a negative feedback mechanism in normal rats. However, renal effects of GLP-1 in heart failure (HF) has not been elucidated. The present study was designed to assess GLP-1-induced diuresis and natriuresis in rats with HF and its interactions with renal nerve activity. METHODS: HF was induced in rats by coronary artery ligation. The direct recording of afferent renal nerve activity (ARNA) with intrapelvic injection of GLP-1 and total renal sympathetic nerve activity (RSNA) with intravenous infusion of GLP-1 were performed. GLP-1 receptor expression in renal pelvis, densely innervated by afferent renal nerve, was assessed by real-time PCR and western blot analysis. In separate group of rats after coronary artery ligation selective afferent renal denervation (A-RDN) was performed by periaxonal application of capsaicin, then intravenous infusion of GLP-1-induced diuresis and natriuresis were evaluated. RESULTS: In HF, compared to sham-operated control; (1) response of increase in ARNA to intrapelvic injection of GLP-1 was enhanced (3.7 ± 0.4 vs. 2.0 ± 0.4 µV s), (2) GLP-1 receptor expression was increased in renal pelvis, (3) response of increase in RSNA to intravenous infusion of GLP-1 was enhanced (132 ± 30% vs. 70 ± 16% of the baseline level), and (4) diuretic and natriuretic responses to intravenous infusion of GLP-1 were blunted (urine flow 53.4 ± 4.3 vs. 78.6 ± 4.4 µl/min/gkw, sodium excretion 7.4 ± 0.8 vs. 10.9 ± 1.0 µEq/min/gkw). A-RDN induced significant increases in diuretic and natriuretic responses to GLP-1 in HF (urine flow 96.0 ± 1.9 vs. 53.4 ± 4.3 µl/min/gkw, sodium excretion 13.6 ± 1.4 vs. 7.4 ± 0.8 µEq/min/gkw). CONCLUSIONS: The excessive activation of neural circuitry involving afferent and efferent renal nerves suppresses diuretic and natriuretic responses to GLP-1 in HF. These pathophysiological responses to GLP-1 might be involved in the interaction between incretin-based medicines and established HF condition. RDN restores diuretic and natriuretic effects of GLP-1 and thus has potential beneficial therapeutic implication for diabetic HF patients.


Assuntos
Capsaicina/administração & dosagem , Diurese/efeitos dos fármacos , Diuréticos/administração & dosagem , Peptídeo 1 Semelhante ao Glucagon/administração & dosagem , Insuficiência Cardíaca/tratamento farmacológico , Rim/efeitos dos fármacos , Rim/inervação , Natriurese/efeitos dos fármacos , Simpatectomia Química , Animais , Modelos Animais de Doenças , Insuficiência Cardíaca/fisiopatologia , Infusões Intravenosas , Masculino , Ratos Sprague-Dawley
7.
Nitric Oxide ; 94: 54-62, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31654775

RESUMO

Activation of renin-angiotensin- system, nitric oxide (NO•) bioavailability and subsequent sympathoexcitation plays a pivotal role in the pathogenesis of many cardiovascular diseases, including hypertension. Previously we have shown increased protein expression of PIN (a protein inhibitor of nNOS: neuronal nitric oxide synthase, known to dissociate nNOS dimers into monomers) with concomitantly reduced levels of catalytically active dimers of nNOS in the PVN of rats with heart failure. To elucidate the molecular mechanism by which Angiotensin II (Ang II) increases PIN expression, we used Sprague-Dawley rats (250-300 g) subjected to intracerebroventricular infusion of Ang II (20 ng/min, 0.5 µl/h) or saline as vehicle (Veh) for 14 days through osmotic mini-pumps and NG108-15 hybrid neuronal cell line treated with Ang II as an in vitro model. Ang II infusion significantly increased baseline renal sympathetic nerve activity and mean arterial pressure. Ang II infusion increased the expression of PIN (1.24 ± 0.04* Ang II vs. 0.65 ± 0.07 Veh) with a concomitant 50% decrease in dimeric nNOS and PIN-Ub conjugates (0.73 ± 0.04* Ang II vs. 1.00 ± 0.03 Veh) in the PVN. Substrate-dependent ligase assay in cells transfected with pCMV-(HA-Ub)8 vector revealed a reduction of HA-Ub-PIN conjugates after Ang II and a proteasome inhibitor, Lactacystin (LC), treatment (4.5 ± 0.7* LC Ang II vs. 9.2 ± 2.5 LC). TUBE (Tandem Ubiquitin-Binding Entities) assay showed decrease PIN-Ub conjugates in Ang II-treated cells (0.82 ± 0.12* LC Ang II vs. 1.21 ± 0.06 LC) while AT1R blocker, Losartan (Los) treatment diminished the Ang II-mediated stabilization of PIN (1.21 ± 0.07 LC Los vs. 1.16 ± 0.04* LC Ang II Los). Taken together, our studies suggest that increased central levels of Ang II contribute to the enhanced expression of PIN leading to reduced expression of the dimeric form of nNOS, thus diminishing the inhibitory action of NO• on pre-autonomic neurons in the PVN resulting in increased sympathetic outflow.


Assuntos
Angiotensina II/administração & dosagem , Hipertensão/induzido quimicamente , Angiotensina II/farmacologia , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Hipertensão/metabolismo , Infusões Intraventriculares , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo I/antagonistas & inibidores , Óxido Nítrico Sintase Tipo I/metabolismo , Ratos , Ratos Sprague-Dawley , Células Tumorais Cultivadas
8.
Sci Rep ; 9(1): 12986, 2019 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537818

RESUMO

Glucagon-like peptide-1 receptor (GLP-1R) agonists, widely used to treat type 2 diabetes, reduce blood pressure (BP) in hypertensive patients. Whether this action involves central mechanisms is unknown. We here report that repeated lateral ventricular (LV) injection of GLP-1R agonist, liraglutide, once daily for 15 days counteracted the development of hypertension in spontaneously hypertensive rats (SHR). In parallel, it suppressed urinary norepinephrine excretion, and induced c-Fos expressions in the area postrema (AP) and nucleus tractus solitarius (NTS) of brainstem including the NTS neurons immunoreactive to dopamine beta-hydroxylase (DBH). Acute administration of liraglutide into fourth ventricle, the area with easy access to the AP and NTS, transiently decreased BP in SHR and this effect was attenuated after lesion of NTS DBH neurons with anti-DBH conjugated to saporin (anti-DBH-SAP). In anti-DBH-SAP injected SHR, the antihypertensive effect of repeated LV injection of liraglutide for 14 days was also attenuated. These findings demonstrate that the central GLP-1R signaling via NTS DBH neurons counteracts the development of hypertension in SHR, accompanied by attenuated sympathetic nerve activity.


Assuntos
Tronco Encefálico/metabolismo , Neurônios Dopaminérgicos/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Hipertensão/metabolismo , Transdução de Sinais , Animais , Tronco Encefálico/patologia , Dopamina beta-Hidroxilase/metabolismo , Neurônios Dopaminérgicos/patologia , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Hipertensão/induzido quimicamente , Hipertensão/patologia , Liraglutida/efeitos adversos , Liraglutida/farmacologia , Masculino , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY
9.
Am J Physiol Renal Physiol ; 317(4): F1010-F1021, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31390233

RESUMO

Glucagon-like peptide-1 (GLP-1), an incretin hormone, has diuretic and natriuretic effects. The present study was designed to explore the possible underlying mechanisms for the diuretic and natriuretic effects of GLP-1 via renal nerves in rats. Immunohistochemistry revealed that GLP-1 receptors were avidly expressed in the pelvic wall, the wall being adjacent to afferent renal nerves immunoreactive to calcitonin gene-related peptide, which is the dominant neurotransmitter for renal afferents. GLP-1 (3 µM) infused into the left renal pelvis increased ipsilateral afferent renal nerve activity (110.0 ± 15.6% of basal value). Intravenous infusion of GLP-1 (1 µg·kg-1·min-1) for 30 min increased renal sympathetic nerve activity (RSNA). After the distal end of the renal nerve was cut to eliminate the afferent signal, the increase in efferent renal nerve activity during intravenous infusion of GLP-1 was diminished compared with the increase in total RSNA (17.0 ± 9.0% vs. 68.1 ± 20.0% of the basal value). Diuretic and natriuretic responses to intravenous infusion of GLP-1 were enhanced by total renal denervation (T-RDN) with acute surgical cutting of the renal nerves. Selective afferent renal nerve denervation (A-RDN) was performed by bilateral perivascular application of capsaicin on the renal nerves. Similar to T-RDN, A-RDN enhanced diuretic and natriuretic responses to GLP-1. Urine flow and Na+ excretion responses to GLP-1 were not significantly different between T-RDN and A-RDN groups. These results indicate that the diuretic and natriuretic effects of GLP-1 are partly governed via activation of afferent renal nerves by GLP-1 acting on sensory nerve fibers within the pelvis of the kidney.


Assuntos
Vias Aferentes/efeitos dos fármacos , Diurese/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Rim/efeitos dos fármacos , Rim/inervação , Natriurese/efeitos dos fármacos , Animais , Peptídeo Relacionado com Gene de Calcitonina/fisiologia , Denervação , Peptídeo 1 Semelhante ao Glucagon/biossíntese , Receptor do Peptídeo Semelhante ao Glucagon 1/biossíntese , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Receptor do Peptídeo Semelhante ao Glucagon 1/imunologia , Células HEK293 , Humanos , Pelve Renal/efeitos dos fármacos , Pelve Renal/inervação , Masculino , Ratos , Ratos Sprague-Dawley , Circulação Renal/efeitos dos fármacos , Sódio/urina , Sistema Nervoso Simpático/efeitos dos fármacos , Urodinâmica/efeitos dos fármacos
10.
Adv Physiol Educ ; 43(2): 175-179, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30998107

RESUMO

Scientific advocacy and outreach programs are encouraged to increase public understanding of scientific knowledge and generate interest in science, technology, engineering, and mathematics (STEM) careers. However, evaluation of these events' effectiveness is difficult and somewhat rare. This study's purpose was to better understand how effective an established physiology-based outreach program was in generating interest in STEM careers, while simultaneously providing information that can be used to increase the effectiveness of future events. We partnered with a private school located in Omaha, Nebraska, where 64-80 students participated in 3 h of physiology-based activities presented by volunteers from the University of Nebraska Medical Center. The event included a brief presentation of the eye, sensory, heart, and lung systems, followed by hands-on demonstrations and activities. Each session concluded with 15 min of questions and answers (Q&A), where students were encouraged to engage the volunteers in inquiries about what they just learned, career-related questions, or any topic of their choosing. Each Q&A session was audio recorded and evaluated using thematic analysis to identify patterns in the Q&A data. Two major themes of questions were identified: 1) scientific content (animal circulatory systems and how organs are affected by disease or stimulus); and 2) career-related content, including typical day-to-day activities of a scientist and the volunteers' satisfaction with a scientific career. We conclude that hands-on physiology-based learning opportunities are effective in generating short-term interest in STEM content and careers. The results of this study will also facilitate informed modification of event content to better suit student's interests.


Assuntos
Escolha da Profissão , Relações Comunidade-Instituição , Matemática/educação , Fisiologia/educação , Humanos , Matemática/métodos , Motivação , Fisiologia/métodos , Estudantes/psicologia
11.
Nitric Oxide ; 87: 73-82, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30878404

RESUMO

Exercise training (ExT) is an established non-pharmacological therapy that improves the health and quality of life in patients with chronic heart failure (CHF). Exaggerated sympathetic drive characterizes CHF due to an imbalance of the autonomic nervous system. Neuronal nitric oxide synthase (nNOS) in the paraventricular nucleus (PVN) produce nitric oxide (NO•), which is known to regulate the sympathetic tone. Previously we have shown that during CHF, the catalytically active dimeric form of nNOS is significantly decreased with a concurrent increase in protein inhibitor of nNOS (PIN) expression, a protein that dissociates dimeric nNOS to monomers and facilitates its degradation. Dimerization of nNOS also requires (6R)-5,6,7,8-tetrahydrobiopterin (BH4) for stability and activity. Previously, we have shown that ExT improves NO-mediated sympathetic inhibition in the PVN; however, the molecular mechanism remains elusive. We hypothesized; ExT restores the sympathetic drive by increasing the levels and catalytically active form of nNOS by abrogating changes in the PIN in the PVN of CHF rats. CHF was induced in adult male Sprague-Dawley rats by coronary artery ligation, which reliably mimics CHF in patients with myocardial infarction. After 4 weeks of surgery, Sham and CHF rats were subjected to 3 weeks of progressive treadmill exercise. ExT significantly (p < 0.05) decreased PIN expression and increased dimer/monomer ratio of nNOS in the PVN of rats with CHF. Moreover, we found decreased GTP cyclohydrolase 1(GCH1) expression: a rate-limiting enzyme for BH4 biosynthesis in the PVN of CHF rats suggesting that perhaps reduced BH4 availability may also contribute to decreased nNOS dimers. Interestingly, CHF induced decrease in GCH1 expression was increased with ExT. Our findings revealed that ExT rectified decreased PIN and GCH1 expression and increased dimer/monomer ratio of nNOS in the PVN, which may lead to increase NO• bioavailability resulting in amelioration of activated sympathetic drive during CHF.


Assuntos
Insuficiência Cardíaca/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Condicionamento Físico Animal/fisiologia , Multimerização Proteica/fisiologia , Animais , Biopterinas/análogos & derivados , Biopterinas/metabolismo , Doença Crônica , Dineínas do Citoplasma/metabolismo , GTP Cicloidrolase/metabolismo , Masculino , Ratos Sprague-Dawley
12.
Nanomedicine ; 14(3): 781-788, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29278747

RESUMO

There are multiple challenges for neuropharmacology in the future. Undoubtedly, one of the greatest challenges is the development of strategies for pharmacological targeting of specific brain regions for treatment of diseases. GABA is the main inhibitory neurotransmitter in the central nervous system, and dysfunction of GABAergic mechanisms is associated with different neurological conditions. Liposomes are lipid vesicles that are able to encapsulate chemical compounds and are used for chronic drug delivery. This short review reports our experience with the development of liposomes for encapsulation and chronic delivery of GABA to sites within the brain. Directions for future research regarding the efficacy and practical use of GABA-containing liposomes for extended periods of time as well as understanding and targeting neurological conditions are discussed.


Assuntos
Sistemas de Liberação de Medicamentos , Lipossomos/administração & dosagem , Doenças do Sistema Nervoso/terapia , Ácido gama-Aminobutírico/administração & dosagem , Animais , Humanos , Lipossomos/química , Ácido gama-Aminobutírico/química
13.
Am J Physiol Heart Circ Physiol ; 312(5): H968-H979, 2017 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-28283551

RESUMO

An activated renin-angiotensin system (RAS) within the central nervous system has been implicated in sympathoexcitation during various disease conditions including congestive heart failure (CHF). In particular, activation of the RAS in the paraventricular nucleus (PVN) of the hypothalamus has been recognized to augment sympathoexcitation in CHF. We observed a 2.6-fold increase in angiotensinogen (AGT) in the PVN of CHF. To elucidate the molecular mechanism for increased expression of AGT, we performed in silico analysis of the 3'-untranslated region (3'-UTR) of AGT and found a potential binding site for microRNA (miR)-133a. We hypothesized that decreased miR-133a might contribute to increased AGT in the PVN of CHF rats. Overexpression of miR-133a in NG108 cells resulted in 1.4- and 1.5-fold decreases in AGT and angiotensin type II (ANG II) type 1 receptor (AT1R) mRNA levels, respectively. A luciferase reporter assay performed on NG108 cells confirmed miR-133a binding to the 3'-UTR of AGT. Consistent with these in vitro data, we observed a 1.9-fold decrease in miR-133a expression with a concomitant increase in AGT and AT1R expression within the PVN of CHF rats. Furthermore, restoring the levels of miR-133a within the PVN of CHF rats with viral transduction resulted in a significant reduction of AGT (1.4-fold) and AT1R (1.5-fold) levels with a concomitant decrease in basal renal sympathetic nerve activity (RSNA). Restoration of miR-133a also abrogated the enhanced RSNA responses to microinjected ANG II within the PVN of CHF rats. These results reveal a novel and potentially unique role for miR-133a in the regulation of ANG II within the PVN of CHF rats, which may potentially contribute to the commonly observed sympathoexcitation in CHF.NEW & NOTEWORTHY Angiotensinogen (AGT) expression is upregulated in the paraventricular nucleus of the hypothalamus through posttranscriptional mechanism interceded by microRNA-133a in heart failure. Understanding the mechanism of increased expression of AGT in pathological conditions leading to increased sympathoexcitation may provide the basis for the possible development of new therapeutic agents with enhanced specificity.


Assuntos
Insuficiência Cardíaca/genética , MicroRNAs/genética , Sistema Renina-Angiotensina/genética , Regiões 3' não Traduzidas , Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Angiotensinogênio/metabolismo , Animais , Linhagem Celular Tumoral , Insuficiência Cardíaca/fisiopatologia , Rim/inervação , Losartan/uso terapêutico , Masculino , MicroRNAs/biossíntese , Núcleo Hipotalâmico Paraventricular/fisiopatologia , Ratos , Ratos Sprague-Dawley , Receptor Tipo 1 de Angiotensina/biossíntese , Receptor Tipo 1 de Angiotensina/genética
14.
Expert Opin Ther Targets ; 21(1): 11-22, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27885874

RESUMO

INTRODUCTION: Nitric oxide (NO) synthesized via neuronal nitric oxide synthase (nNOS) plays a significant role in regulation/modulation of autonomic control of circulation. Various pathological states are associated with diminished nNOS expression and blunted autonomic effects of NO in the central nervous system (CNS) including heart failure, hypertension, diabetes mellitus, chronic renal failure etc. Therefore, elucidation of the molecular mechanism/s involved in dysregulation of nNOS is essential to understand the pathogenesis of increased sympathoexcitation in these diseased states. Areas covered: nNOS is a highly regulated enzyme, being regulated at transcriptional and posttranslational levels via protein-protein interactions and modifications viz. phosphorylation, ubiquitination, and sumoylation. The enzyme activity of nNOS also depends on the optimal concentration of substrate, cofactors and association with regulatory proteins. This review focuses on the posttranslational regulation of nNOS in the context of normal and diseased states within the CNS. Expert opinion: Gaining insight into the mechanism/s involved in the regulation of nNOS would provide novel strategies for manipulating nNOS directed therapeutic modalities in the future, including catalytically active dimer stabilization and protein-protein interactions with intracellular protein effectors. Ultimately, this is expected to provide tools to improve autonomic dysregulation in various diseases such as heart failure, hypertension, and diabetes.


Assuntos
Óxido Nítrico Sintase Tipo I/metabolismo , Óxido Nítrico/metabolismo , Sistema Nervoso Simpático/metabolismo , Animais , Sistema Nervoso Autônomo/fisiologia , Diabetes Mellitus/fisiopatologia , Insuficiência Cardíaca/fisiopatologia , Humanos , Hipertensão/fisiopatologia , Processamento de Proteína Pós-Traducional/fisiologia
15.
Circ Heart Fail ; 9(11)2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27810863

RESUMO

BACKGROUND: Increased sympathetic outflow is a major contributor to the progression of chronic heart failure (CHF). Potentiation of glutamatergic tone has been causally related to the sympathoexcitation in CHF. Specifically, an increase in the N-methyl-d-aspartate-type 1 receptor (NMDA-NR1) expression within the paraventricular nucleus (PVN) is critically linked to the increased sympathoexcitation during CHF. However, the molecular mechanism(s) for the upregulation of NMDA-NR1 remains unexplored. We hypothesized that hypoxia via hypoxia-inducible factor 1α (HIF-1α) might contribute to the augmentation of the NMDA-NR1-mediated sympathoexcitatory responses from the PVN in CHF. METHODS AND RESULTS: Immunohistochemistry staining, mRNA, and protein for hypoxia-inducible factor 1α were upregulated within the PVN of left coronary artery-ligated CHF rats. In neuronal cell line (NG108-15) in vitro, hypoxia caused a significant increase in mRNA and protein for HIF-1α (2-fold) with the concomitant increase in NMDA-NR1 mRNA, protein levels, and glutamate-induced Ca+ influx. Chromatin immunoprecipitation assay identified HIF-1α binding to NMDA-NR1 promoter during hypoxia. Silencing of HIF-1α in NG108 cells leads to a significant decrease in expression of NMDA-NR1, suggesting that expression of HIF-1α is necessary for the upregulation of NMDA-NR1. Consistent with these observations, HIF-1α silencing within the PVN abrogated the increased basal sympathetic tone and sympathoexcitatory responses to microinjection of NMDA in the PVN of rats with CHF. CONCLUSIONS: These results uncover a critical role for HIF-1 in the upregulation of NMDA-NR1 to mediate sympathoexcitation in CHF. We conclude that subtle hypoxia within the PVN may act as a metabolic cue to modulate sympathoexcitation during CHF.


Assuntos
Insuficiência Cardíaca/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Hipóxia/genética , Neurônios/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , RNA Mensageiro/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Sistema Nervoso Simpático/fisiopatologia , Animais , Linhagem Celular , Imunoprecipitação da Cromatina , Doença Crônica , Modelos Animais de Doenças , Progressão da Doença , Agonistas de Aminoácidos Excitatórios/farmacologia , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imuno-Histoquímica , Técnicas In Vitro , Masculino , N-Metilaspartato/farmacologia , Núcleo Hipotalâmico Paraventricular/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo , Sistema Nervoso Simpático/efeitos dos fármacos , Regulação para Cima
16.
J Neurosci Methods ; 273: 55-63, 2016 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-27523033

RESUMO

BACKGROUND: Liposomes are concentric lipid vesicles that allow a sustained release of entrapped substances. GABA (γ-aminobutyric acid) is the most prevalent inhibitory neurotransmitter in the central nervous system. NEW METHOD: Using GABA-containing liposomes (GL) prepared by the freeze-thawing method, we determined the effect of sustained release of GABA on expression of neuronal nitric oxide synthase (nNOS) and GABAA receptor (GABAAR) in an in vitro neuronal model. RESULTS: Neuronal cell line NG108-15 treated with different doses of GL during 24h showed an increase in expression of GABAAR (54 and 50% with 10 and 20ng doses, respectively) and nNOS (138, 157 and 165% with 20, 50 and 100ng doses, respectively) compared with cells treated with empty liposomes (EL). Additionally, cells treated with 50ng of GL showed an increase in GABAAR (23%) after 1h followed by an increase in nNOS (55, 46 and 55%) at 8, 12 and 24h time points, respectively. Immunofluorescence experiments confirmed an increase in nNOS (134%) and basal intracellular levels of nitric oxide (84%) after GL treatment. Further, treatment of cells with GL showed a decrease in expression of a protein inhibitor of nNOS (PIN) (26, 66 and 57% with 20, 50 and 100ng doses respectively) compared with control. COMPARISON WITH EXISTING METHODS: This is first demonstration for the development of GL that allows sustained slow release of this neurotransmitter. CONCLUSION: These results suggest that a slow release of GABA can change the expression of nNOS possibly via alteration in PIN levels in neuronal cells.


Assuntos
Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Lipossomos/administração & dosagem , Óxido Nítrico Sintase Tipo I/metabolismo , Ácido gama-Aminobutírico/administração & dosagem , Animais , Proteína de Ligação a CREB/metabolismo , Linhagem Celular Tumoral , Dineínas do Citoplasma/metabolismo , Relação Dose-Resposta a Droga , Sistemas de Liberação de Medicamentos , Ácido Glutâmico/farmacologia , Lipossomos/farmacologia , Camundongos , Neuroblastoma/patologia , Nitritos/metabolismo , RNA Interferente Pequeno/farmacologia , Ratos , Receptores de GABA-A/metabolismo , Fatores de Tempo , Ácido gama-Aminobutírico/farmacologia
17.
Diabetes ; 65(10): 3075-90, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27411382

RESUMO

MicroRNAs (miRNAs) have a fundamental role in diabetic heart failure. The cardioprotective miRNA-133a (miR-133a) is downregulated, and contractility is decreased in diabetic hearts. Norepinephrine (NE) is a key catecholamine that stimulates contractility by activating ß-adrenergic receptors (ß-AR). NE is synthesized from tyrosine by the rate-limiting enzyme, tyrosine hydroxylase (TH), and tyrosine is catabolized by tyrosine aminotransferase (TAT). However, the cross talk/link between TAT and TH in the heart is unclear. To determine whether miR-133a plays a role in the cross talk between TH and TAT and regulates contractility by influencing NE biosynthesis and/or ß-AR levels in diabetic hearts, Sprague-Dawley rats and miR-133a transgenic (miR-133aTg) mice were injected with streptozotocin to induce diabetes. The diabetic rats were then treated with miR-133a mimic or scrambled miRNA. Our results revealed that miR-133a mimic treatment improved the contractility of the diabetic rat's heart concomitant with upregulation of TH, cardiac NE, ß-AR, and downregulation of TAT and plasma levels of NE. In miR-133aTg mice, cardiac-specific miR-133a overexpression prevented upregulation of TAT and suppression of TH in the heart after streptozotocin was administered. Moreover, miR-133a overexpression in CATH.a neuronal cells suppressed TAT with concomitant upregulation of TH, whereas knockdown and overexpression of TAT demonstrated that TAT inhibited TH. Luciferase reporter assay confirmed that miR-133a targets TAT. In conclusion, miR-133a controls the contractility of diabetic hearts by targeting TAT, regulating NE biosynthesis, and consequently, ß-AR and cardiac function.


Assuntos
MicroRNAs/metabolismo , Contração Miocárdica/fisiologia , Miocárdio/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Tirosina Transaminase/metabolismo , Animais , Western Blotting , Diabetes Mellitus Experimental/metabolismo , Células HEK293 , Hemodinâmica/fisiologia , Humanos , Imuno-Histoquímica , Masculino , Camundongos Transgênicos , MicroRNAs/genética , MicroRNAs/fisiologia , Contração Miocárdica/genética , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/fisiologia , Norepinefrina/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Adrenérgicos beta/genética , Receptores Adrenérgicos beta/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tirosina 3-Mono-Oxigenase/genética , Tirosina Transaminase/genética
18.
Am J Physiol Heart Circ Physiol ; 311(2): H337-46, 2016 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-27288440

RESUMO

Chronic activation of the sympathetic drive contributes to cardiac remodeling and dysfunction during chronic heart failure (HF). The present study was undertaken to assess whether renal denervation (RDN) would abrogate the sympathoexcitation in HF and ameliorate the adrenergic dysfunction and cardiac damage. Ligation of the left coronary artery was used to induce HF in Sprague-Dawley rats. Four weeks after surgery, RDN was performed, 1 wk before the final measurements. At the end of the protocol, cardiac function was assessed by measuring ventricular hemodynamics. Rats with HF had an average infarct area >30% of the left ventricle and left ventricular end-diastolic pressure (LVEDP) >20 mmHg. ß1- and ß2-adrenoceptor proteins in the left ventricle were reduced by 37 and 49%, respectively, in the rats with HF. RDN lowered elevated levels of urinary excretion of norepinephrine and brain natriuretic peptide levels in the hearts of rats with HF. RDN also decreased LVEDP to 10 mmHg and improved basal dP/dt to within the normal range in rats with HF. RDN blunted loss of ß1-adrenoceptor (by 47%) and ß2-adrenoceptor (by 100%) protein expression and improved isoproterenol (0.5 µg/kg)-induced increase in +dP/dt (by 71%) and -dP/dt (by 62%) in rats with HF. RDN also attenuated the increase in collagen 1 expression in the left ventricles of rats with HF. These findings demonstrate that RDN initiated in chronic HF condition improves cardiac function mediated by adrenergic agonist and blunts ß-adrenoceptor expression loss, providing mechanistic insights for RDN-induced improvements in cardiac function in the HF condition.


Assuntos
Insuficiência Cardíaca/fisiopatologia , Ventrículos do Coração/fisiopatologia , Artéria Renal/inervação , Simpatectomia , Agonistas Adrenérgicos beta/farmacologia , Animais , Doença Crônica , Colágeno Tipo I/metabolismo , Vasos Coronários/cirurgia , Coração/efeitos dos fármacos , Insuficiência Cardíaca/etiologia , Ventrículos do Coração/metabolismo , Isoproterenol/farmacologia , Córtex Renal/metabolismo , Ligadura , Masculino , Infarto do Miocárdio/complicações , Peptídeo Natriurético Encefálico/metabolismo , Norepinefrina/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Adrenérgicos beta 1/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Sódio/urina
19.
Hypertension ; 68(1): 175-84, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27185748

RESUMO

Renal denervation (RDN) has been postulated to reduce sympathetic drive during heart failure (HF), but the central mechanisms are not completely understood. The purpose of the present study was to assess the contribution of neuronal nitric oxide synthase (nNOS) within the paraventricular nucleus (PVN) in modulating sympathetic outflow in rats with HF that underwent RDN. HF was induced in rats by ligation of the left coronary artery. Four weeks after surgery, bilateral RDN was performed. Rats with HF had an increase in FosB-positive cells in the PVN with a concomitant increase in urinary excretion of norepinephrine, and both of these parameters were ameliorated after RDN. nNOS-positive cells immunostaining, diaphorase staining, and nNOS protein expression were significantly decreased in the PVN of HF rats, findings that were ameliorated by RDN. Microinjection of nNOS inhibitor N(G)-monomethyl l-arginine into the PVN resulted in a blunted increase in lumbar sympathetic nerve activity (11±2% versus 24±2%) in HF than in sham group. This response was normalized after RDN. Stimulation of afferent renal nerves produced a greater activation of PVN neurons in rats with HF. Afferent renal nerve stimulation elicited a greater increase in lumbar sympathetic nerve activity in rats with HF than in sham rats (45±5% versus 22±2%). These results suggest that intact renal nerves contribute to the reduction of nNOS in the PVN, resulting in the activation of the neurons in the PVN of rats with HF. RDN restores nNOS and thus attenuates the sympathoexcitation commonly observed in HF.


Assuntos
Insuficiência Cardíaca/metabolismo , Rim/inervação , Óxido Nítrico Sintase Tipo I/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Sistema Nervoso Simpático/fisiopatologia , Animais , Denervação Autônoma/métodos , Modelos Animais de Doenças , Insuficiência Cardíaca/fisiopatologia , Rim/cirurgia , Masculino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Valores de Referência
20.
Oxid Med Cell Longev ; 2016: 5868913, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26989452

RESUMO

Exercise training (ExT) is currently being used as a nonpharmacological strategy to improve cardiac function in diabetic patients. However, the molecular mechanism(s) underlying its beneficial effects remains poorly understood. Oxidative stress is known to play a key role in the pathogenesis of diabetic cardiomyopathy and one of the enzyme systems that produce reactive oxygen species is NADH/NADPH oxidase. The goal of this study was to investigate the effect of streptozotocin- (STZ-) induced diabetes on expression of p47(phox) and p67(phox), key regulatory subunits of NADPH oxidase, in cardiac tissues and determine whether ExT can attenuate these changes. Four weeks after STZ treatment, expression of p47(phox) and p67(phox) increased 2.3-fold and 1.6-fold, respectively, in left ventricles of diabetic rats and these increases were attenuated with three weeks of ExT, initiated 1 week after onset of diabetes. In atrial tissues, there was increased expression of p47(phox) (74%), which was decreased by ExT in diabetic rats. Furthermore, increased collagen III levels in diabetic hearts (52%) were significantly reduced by ExT. Taken together, ExT attenuates the increased expression of p47(phox) and p67(phox) in the hearts of diabetic rats which could be an underlying mechanism for improving intracardiac matrix and thus cardiac function and prevent cardiac remodeling in diabetic cardiomyopathy.


Assuntos
Diabetes Mellitus Experimental/enzimologia , Cardiomiopatias Diabéticas/enzimologia , Regulação Enzimológica da Expressão Gênica , Miocárdio/enzimologia , NADH NADPH Oxirredutases/biossíntese , NADPH Oxidases/biossíntese , Condicionamento Físico Animal , Regulação para Cima , Animais , Diabetes Mellitus Experimental/patologia , Cardiomiopatias Diabéticas/patologia , Masculino , Miocárdio/patologia , Ratos , Ratos Sprague-Dawley
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