Lucas Rios Drummond

The benefits of endurance training in cardiomyocyte function in hypertensive rats are reversed within four weeks of detraining

The aim of the present study was to verify the effects of low-intensity endurance training and detraining on the mechanical and molecular properties of cardiomyocytes from spontaneously hypertensive rats (SHRs). Male SHRs and normotensive control Wistar rats at 16-weeks of age were randomly divided into eight groups of eight animals: NC8 and HC8 (normotensive and hypertensive control for 8weeks); NT8 and HT8 (normotensive and hypertensive trained at 50-60% of maximal exercise capacity for 8weeks); NC12 and HC12 (normotensive and hypertensive control for 12weeks); NDT and HDT (normotensive and hypertensive trained for 8weeks and detrained for 4weeks). The total exercise time until fatigue (TTF) was determined by a maximal exercise capacity test. Resting heart rate (RHR) and systolic arterial pressure (SAP) were measured. After the treatments, animals were killed by cervical dislocation and left ventricular myocytes were isolated by enzymatic dispersion. Isolated cells were used to determine intracellular global Ca(2+) ([Ca(2+)]i) transient and cardiomyocyte contractility (1Hz; ~25°C). [Ca(2+)]i regulatory proteins were measured by Western blot, and the markers of pathologic cardiac hypertrophy by quantitative real-time polymerase chain reaction (q-RT-PCR). Exercise training augmented the TTF (NC8, 11.4±1.5min vs. NT8, 22.5±1.4min; HC8, 11.7±1.4min vs. HT8, 24.5±1.3min; P<0.05), reduced RHR (NT8initial, 340±8bpm vs. NT8final, 322±10bpm; HT8initial, 369±8bpm vs. HT8final, 344±10bpm; P<0.05), and SBP in SHR animals (HC8, 178±3mmHg vs. HT8, 161±4mmHg; P<0.05). HC8 rats showed a slower [Ca(2+)]i transient (Tpeak, 83.7±1.8ms vs. 71.7±2.4ms; T50%decay, 284.0±4.3ms vs. 264.0±4.1ms; P<0.05) and cell contractility (Vshortening, 86.1±6.7μm/s vs. 118.6±6.7μm/s; Vrelengthening, 57.5±7.4μm/s vs. 101.3±7.4μm/s; P<0.05), and higher expression of ANF (300%; P<0.05), skeletal α-actin (250%; P<0.05) and a decreased α/β-MHC ratio (70%; P<0.05) compared to NC8. Exercise training increased [Ca(2+)]i transient (NC8, 2.39±0.06F/F0 vs. NT8, 2.72±0.06F/F0; HC8, 2.28±0.05F/F0 vs. HT8, 2.82±0.05F/F0; P<0.05), and cell contractility (NC8, 7.4±0.3% vs. NT8, 8.4±0.3%; HC8, 6.8±0.3% vs. HT8, 7.8±0.3%; P<0.05). Furthermore, exercise normalized the expression of ANF, skeletal α-actin, and the α/β-MHC ratio in HT8 rats, augmented the expression of SERCA2a (NC8, 0.93±0.15 vs. NT8, 1.49±0.14; HC8, 0.83±0.13 vs. HT8, 1.32±0.14; P<0.05) and PLBser16 (NC8, 0.89±0.18 vs. NT8, 1.23±0.17; HC8, 0.77±0.17 vs. HT8, 1.32±0.16; P<0.05), and reduced PLBt/SERCA2a (NC8, 1.21±0.19 vs. NT8, 0.50±0.21; HC8, 1.38±0.17 vs. HT8, 0.66±0.21; P<0.05). However, all these adaptations returned to control values within 4weeks of detraining in both SHR and normotensive control animals. In conclusion, low-intensity endurance training induces positive benefits to left ventricular myocyte mechanical and molecular properties, which are reversed within 4weeks of detraining.

Resistance training regulates cardiac function through modulation of miRNA-214.

Aims: To determine the effects of resistance training (RT) on the expression of microRNA (miRNA)-214 and its target in sarcoplasmic reticulum Ca2+-ATPase (SERCA2a), and on the morphological and mechanical properties of isolated left ventricular myocytes. Main methods: Male Wistar rats were divided into two groups (n = 7/group): Control (CO) or trained (TR). The exercise-training protocol consisted of: 4 × 12 bouts, 5×/week during 8 weeks, with 80% of one repetition maximum. Key findings: RT increased the left ventricular myocyte width by 15% and volume by 12%, compared with control animals (p < 0.05). The time to half relaxation and time to peak were 8.4% and 4.4% lower, respectively, in cells from TR group as compared to CO group (p < 0.05). RT decreased miRNA-214 level by 18.5% while its target SERCA2a expression were 18.5% higher (p < 0.05). Significance: Our findings showed that RT increases single left ventricular myocyte dimensions and also leads to faster cell contraction and relaxation. These mechanical adaptations may be related to the augmented expression of SERCA2a which, in turn, may be associated with the epigenetic modification of decreased miRNA-214 expression.

Thermoregulatory responses in exercising rats: methodological aspects and relevance to human physiology

Rats are used worldwide in experiments that aim to investigate the physiological responses induced by a physical exercise session. Changes in body temperature regulation, which may affect both the performance and the health of exercising rats, are evident among these physiological responses. Despite the universal use of rats in biomedical research involving exercise, investigators often overlook important methodological issues that hamper the accurate measurement of clear thermoregulatory responses. Moreover, much debate exists regarding whether the outcome of rat experiments can be extrapolated to human physiology, including thermal physiology. Herein, we described the impact of different exercise intensities, durations and protocols and environmental conditions on running-induced thermoregulatory changes. We focused on treadmill running because this type of exercise allows for precise control of the exercise intensity and the measurement of autonomic thermoeffectors associated with heat production and loss. Some methodological issues regarding rat experiments, such as the sites for body temperature measurements and the time of day at which experiments are performed, were also discussed. In addition, we analyzed the influence of a high body surface area-to-mass ratio and limited evaporative cooling on the exercise-induced thermoregulatory responses of running rats and then compared these responses in rats to those observed in humans. Collectively, the data presented in this review represent a reference source for investigators interested in studying exercise thermoregulation in rats. In addition, the present data indicate that the thermoregulatory responses of exercising rats can be extrapolated, with some important limitations, to human thermal physiology.

Regional effects of low-intensity endurance training on structural and mechanical properties of rat ventricular myocytes

We tested the effects of low-intensity endurance training (LIET) on the structural and mechanical properties of right (RV) and left ventricular (LV) myocytes. Male Wistar rats (4 mo old) were randomly divided into control (C, n = 7) and trained (T, n = 7, treadmill running at 50-60% of maximal running speed for 8 wk) groups. Isolated ventricular myocyte dimensions, contractility, Ca(2+) transients {intracellular Ca(2+) concentration ([Ca(2+)]i)}, and ventricular [Ca(2+)]i regulatory proteins were measured. LIET augmented cell length (C, 152.5 ± 2.0 μm vs. T, 162.2 ± 2.1 μm; P < 0.05) and volume (C, 5,162 ± 131 μm(3) vs. T, 5,506 ± 132 μm(3); P < 0.05) in the LV but not in the RV. LIET increased cell shortening (C, 7.5 ± 0.3% vs. T, 8.6 ± 0.3%; P < 0.05), the [Ca(2+)]i transient amplitude (C, 2.49 ± 0.06 F/F0 vs. T, 2.82 ± 0.06 F/F0; P < 0.05), the expression of sarcoplasmic reticulum Ca(2+)-ATPase 2a (C, 1.07 ± 0.13 vs. T, 1.59 ± 0.12; P < 0.05), and the levels of phosphorylated phospholamban at serine 16 (C, 0.99 ± 0.11 vs. T, 1.34 ± 0.10; P < 0.05), and reduced the total phospholamban-to-sarcoplasmic reticulum Ca(2+)-ATPase 2a ratio (C, 1.19 ± 0.15 vs. T, 0.40 ± 0.16; P < 0.05) in the LV without changing such parameters in the RV. In conclusion, LIET affected the structure and improved the mechanical properties of LV but not of RV myocytes in rats, helping to characterize the functional and morphological changes that accompany the endurance training-induced cardiac remodeling.

Effect of exercise training on Ca2+ release units of left ventricular myocytes of spontaneously hypertensive rats

In cardiomyocytes, calcium (Ca2+) release units comprise clusters of intracellular Ca2+ release channels located on the sarcoplasmic reticulum, and hypertension is well established as a cause of defects in calcium release unit function. Our objective was to determine whether endurance exercise training could attenuate the deleterious effects of hypertension on calcium release unit components and Ca2+ sparks in left ventricular myocytes of spontaneously hypertensive rats. Male Wistar and spontaneously hypertensive rats (4 months of age) were divided into 4 groups: normotensive (NC) and hypertensive control (HC), and normotensive (NT) and hypertensive trained (HT) animals (7 rats per group). NC and HC rats were submitted to a low-intensity treadmill running protocol (5 days/week, 1 h/day, 0% grade, and 50-60% of maximal running speed) for 8 weeks. Gene expression of the ryanodine receptor type 2 (RyR2) and FK506 binding protein (FKBP12.6) increased (270%) and decreased (88%), respectively, in HC compared to NC rats. Endurance exercise training reversed these changes by reducing RyR2 (230%) and normalizing FKBP12.6 gene expression (112%). Hypertension also increased the frequency of Ca2+ sparks (HC=7.61±0.26 vs NC=4.79±0.19 per 100 µm/s) and decreased its amplitude (HC=0.260±0.08 vs NC=0.324±0.10 ΔF/F0), full width at half-maximum amplitude (HC=1.05±0.08 vs NC=1.26±0.01 µm), total duration (HC=11.51±0.12 vs NC=14.97±0.24 ms), time to peak (HC=4.84±0.06 vs NC=6.31±0.14 ms), and time constant of decay (HC=8.68±0.12 vs NC=10.21±0.22 ms). These changes were partially reversed in HT rats (frequency of Ca2+ sparks=6.26±0.19 µm/s, amplitude=0.282±0.10 ΔF/F0, full width at half-maximum amplitude=1.14±0.01 µm, total duration=13.34±0.17 ms, time to peak=5.43±0.08 ms, and time constant of decay=9.43±0.15 ms). Endurance exercise training attenuated the deleterious effects of hypertension on calcium release units of left ventricular myocytes.

Attenuation of Ca2+ homeostasis, oxidative stress, and mitochondrial dysfunctions in diabetic rat heart: insulin therapy or aerobic exercise?

We tested the effects of swimming training and insulin therapy, either alone or in combination, on the intracellular calcium ([Ca(2+)]i) homeostasis, oxidative stress, and mitochondrial functions in diabetic rat hearts. Male Wistar rats were separated into control, diabetic, or diabetic plus insulin groups. Type 1 diabetes mellitus was induced by streptozotocin (STZ). Insulin-treated groups received 1 to 4 UI of insulin daily for 8 wk. Each group was divided into sedentary or exercised rats. Trained groups were submitted to swimming (90 min/day, 5 days/wk, 8 wk). [Ca(2+)]i transient in left ventricular myocytes (LVM), oxidative stress in LV tissue, and mitochondrial functions in the heart were assessed. Diabetes reduced the amplitude and prolonged the times to peak and to half decay of the [Ca(2+)]i transient in LVM, increased NADPH oxidase-4 (Nox-4) expression, decreased superoxide dismutase (SOD), and increased carbonyl protein contents in LV tissue. In isolated mitochondria, diabetes increased Ca(2+) uptake, susceptibility to permeability transition pore (MPTP) opening, uncoupling protein-2 (UCP-2) expression, and oxygen consumption but reduced H2O2 release. Swimming training corrected the time course of the [Ca(2+)]i transient, UCP-2 expression, and mitochondrial Ca(2+) uptake. Insulin replacement further normalized [Ca(2+)]i transient amplitude, Nox-4 expression, and carbonyl content. Alongside these benefits, the combination of both therapies restored the LV tissue SOD and mitochondrial O2 consumption, H2O2 release, and MPTP opening. In conclusion, the combination of swimming training with insulin replacement was more effective in attenuating intracellular Ca(2+) disruptions, oxidative stress, and mitochondrial dysfunctions in STZ-induced diabetic rat hearts.

Morfologia e contratilidade em cardiomiócitos de ratos com baixo desempenho para o exercício físico

BACKGROUND Aerobic capacity is essential to physical performance, and low aerobic capacity is related to the triggering of various cardiovascular diseases. OBJECTIVE To compare the morphology and contractility of isolated rat cardiomyocytes with low performance and standard performance for exercise. METHODS Wistar rats with 10 weeks of age underwent a protocol of treadmill running to fatigue, and were divided into two groups: Low Performance (LP) and Standard Performance (SP). Then, the animals were sacrificed, the heart was quickly removed and, by means of enzymatic dissociation, left ventricular cardiomyocytes were isolated. The cell and sarcomeres length and width of cardiomyocytes were measured using an edge detection system. The isolated cardiomyocytes were electrically stimulated at 1 and 3 Hz and cell contraction was measured by registering the change of their length. RESULTS The cell length was shorter in the LP group (157.2 ± 1.3 µm; p < 0.05) compared to SP (161.4 ± 1.3 µm), and the same result was observed for the volume of cardiomyocytes (LP, 25.5 ± 0.4 vs. SP, 26.8 ± 0.4 pL; p < 0.05). The time to peak contraction (LP, 116 ± 1 vs. SP 111 ± 2 ms) and total relaxation (LP, 143 ± 3 vs. SP 232 ± 3 ms) were higher in the LP group. CONCLUSION We conclude that left ventricular myocytes of animals with low performance for exercise are smaller than animals with standard performance. In addition to that, they present losses in contractile capacity.

NITRIC OXIDE AND CA2+ DYNAMICS IN CARDIOMYOCYTES: INFLUENCE OF EXERCISE CAPACITY

Introduccion: La capacidad intrinseca para el ejercicio aerobico esta relacionada con el inotropismo cardiaco. Por otro lado, todavia se desconoce la contribucion del oxido nitrico (ON) como mensajero intracelular sobre la dinamica del Ca2+ en ratones con diferentes capacidades intrinsecas para el ejercicio. Objetivo: Evaluar si el ON modula diferencialmente la variacion transitoria intracelular de Ca2+ y las liberaciones espontaneas de Ca2+ (sparks) en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio. Metodos: Ratones machos Wistar fueron seleccionados como desempeno estandar (DE) y alto desempeno (AD), de acuerdo con la capacidad de ejercicio hasta la fatiga, medida a traves del test de fuerza progresiva en la caminadora o cinta electrica. Los cardiomiocitos de los ratones fueron utilizados para determinar el transito intracelular y sparks de Ca2+ evaluados en microscopio confocal. Para estimar la contribucion del ON fue utilizado un inhibidor de sintesis del ON (L-NAME, 100 µM). Los datos fueron analizados a traves de un ANOVA two-way seguido de un post-test Tukey y presentados como promedios ± EPM. Resultados: Los cardiomiocitos de ratones AD mostraron aumento en la amplitud de la variacion transitoria de Ca2+ en comparacion con los DE. Asi mismo, el L-NAME incremento la amplitud transitoria de Ca2+ solamente en ratones DE. No se encontraron diferencias en la constante del tiempo de decaimiento de la variacion transitoria ( t ) de Ca2+ en cardiomiocitos de ratones DE e AD. Todavia, la administracion de L-NAME mostro una reduccion en el t en cardiomiocitos de ambos los grupos. Cardiomiocitos de ratones AD presentaron menor amplitud y frecuencia de sparks de Ca2+ en comparacion al grupo DE. La administracion de L-NAME incremento la amplitud de sparks de Ca2+ en cardiomiocitos del grupo AD. Conclusion: El ON modula la variacion de Ca2+ y sparks de Ca2+ en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio.

ÓXIDO NÍTRICO Y DINAMICA DE CA2+ EN CARDIOMIOCITOS: INFLUENCIA DE LA CAPACIDAD DE EJERCÍCIO

Introduccion: La capacidad intrinseca para el ejercicio aerobico esta relacionada con el inotropismo cardiaco. Por otro lado, todavia se desconoce la contribucion del oxido nitrico (ON) como mensajero intracelular sobre la dinamica del Ca2+ en ratones con diferentes capacidades intrinsecas para el ejercicio. Objetivo: Evaluar si el ON modula diferencialmente la variacion transitoria intracelular de Ca2+ y las liberaciones espontaneas de Ca2+ (sparks) en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio. Metodos: Ratones machos Wistar fueron seleccionados como desempeno estandar (DE) y alto desempeno (AD), de acuerdo con la capacidad de ejercicio hasta la fatiga, medida a traves del test de fuerza progresiva en la caminadora o cinta electrica. Los cardiomiocitos de los ratones fueron utilizados para determinar el transito intracelular y sparks de Ca2+ evaluados en microscopio confocal. Para estimar la contribucion del ON fue utilizado un inhibidor de sintesis del ON (L-NAME, 100 µM). Los datos fueron analizados a traves de un ANOVA two-way seguido de un post-test Tukey y presentados como promedios ± EPM. Resultados: Los cardiomiocitos de ratones AD mostraron aumento en la amplitud de la variacion transitoria de Ca2+ en comparacion con los DE. Asi mismo, el L-NAME incremento la amplitud transitoria de Ca2+ solamente en ratones DE. No se encontraron diferencias en la constante del tiempo de decaimiento de la variacion transitoria ( t ) de Ca2+ en cardiomiocitos de ratones DE e AD. Todavia, la administracion de L-NAME mostro una reduccion en el t en cardiomiocitos de ambos los grupos. Cardiomiocitos de ratones AD presentaron menor amplitud y frecuencia de sparks de Ca2+ en comparacion al grupo DE. La administracion de L-NAME incremento la amplitud de sparks de Ca2+ en cardiomiocitos del grupo AD. Conclusion: El ON modula la variacion de Ca2+ y sparks de Ca2+ en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio.

ÓXIDO NÍTRICO E DINÂMICA DE CA2+ EM CARDIOMIÓCITOS: INFLUÊNCIA DA CAPACIDADE DE EXERCÍCIO

Introduccion: La capacidad intrinseca para el ejercicio aerobico esta relacionada con el inotropismo cardiaco. Por otro lado, todavia se desconoce la contribucion del oxido nitrico (ON) como mensajero intracelular sobre la dinamica del Ca2+ en ratones con diferentes capacidades intrinsecas para el ejercicio. Objetivo: Evaluar si el ON modula diferencialmente la variacion transitoria intracelular de Ca2+ y las liberaciones espontaneas de Ca2+ (sparks) en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio. Metodos: Ratones machos Wistar fueron seleccionados como desempeno estandar (DE) y alto desempeno (AD), de acuerdo con la capacidad de ejercicio hasta la fatiga, medida a traves del test de fuerza progresiva en la caminadora o cinta electrica. Los cardiomiocitos de los ratones fueron utilizados para determinar el transito intracelular y sparks de Ca2+ evaluados en microscopio confocal. Para estimar la contribucion del ON fue utilizado un inhibidor de sintesis del ON (L-NAME, 100 µM). Los datos fueron analizados a traves de un ANOVA two-way seguido de un post-test Tukey y presentados como promedios ± EPM. Resultados: Los cardiomiocitos de ratones AD mostraron aumento en la amplitud de la variacion transitoria de Ca2+ en comparacion con los DE. Asi mismo, el L-NAME incremento la amplitud transitoria de Ca2+ solamente en ratones DE. No se encontraron diferencias en la constante del tiempo de decaimiento de la variacion transitoria ( t ) de Ca2+ en cardiomiocitos de ratones DE e AD. Todavia, la administracion de L-NAME mostro una reduccion en el t en cardiomiocitos de ambos los grupos. Cardiomiocitos de ratones AD presentaron menor amplitud y frecuencia de sparks de Ca2+ en comparacion al grupo DE. La administracion de L-NAME incremento la amplitud de sparks de Ca2+ en cardiomiocitos del grupo AD. Conclusion: El ON modula la variacion de Ca2+ y sparks de Ca2+ en cardiomiocitos de ratones con diferentes capacidades intrinsecas para el ejercicio.