Serotonergic neurons in the caudal raphe nuclei discharge in association with activity of masticatory muscles

There is a dense serotonergic projection from nucleus raphe pallidus and nucleus raphe obscurus to the trigeminal motor nucleus and serotonin exerts a strong facilitatory action on the trigeminal motoneurons. Some serotonergic neurons in these caudal raphe nuclei increase their discharge during feeding. The objective of the present study was to investigate the possibility that the activity of these serotonergic neurons is related to activity of masticatory muscles. Cats were implanted with microelectrodes and gross electrodes. Caudal raphe single neuron activity, electrocorticographic activity, and splenius, digastric and masseter electromyographic activities were recorded during active behaviors (feeding and grooming), during quiet waking and during sleep. Seven presumed serotonergic neurons were identified. These neurons showed a long duration action potential (>2.0 ms), and discharged slowly (2-7 Hz) and very regularly (interspike interval coefficient of variation <0.3) during quiet waking. The activity of these neurons decreased remarkably during fast wave sleep (78-100%). Six of these neurons showed tonic changes in their activity positively related to digastric and/or masseter muscle activity but not to splenius muscle activity during waking. These data are consistent with the hypothesis that serotonergic neurons in the caudal raphe nuclei play an important role in the control of jaw movements

Behavioral correlates of the activity of serotonergic and non-serotonergic neurons in caudal raphe nuclei

We investigated the behavioral correlates of the activity of serotonergic and non-serotonergic neurons in the nucleus raphe pallidus (NRP) and nucleus raphe obscurus (NRO) of unanesthetized and unrestrained cats. The animals were implanted with electrodes for recording single unit activity, parietal oscillographic activity, and splenius, digastric and masseter electromyographic activities. They were tested along the waking-sleep cycle, during sensory stimulation and during drinking behavior. The discharge of the serotonergic neurons decreased progressively from quiet waking to slow wave sleep and to fast wave sleep. Ten different patterns of relative discharge across the three states were observed for the non-serotonergic neurons. Several non-serotonergic neurons showed cyclic discharge fluctuations related to respiration during one, two or all three states. While serotonergic neurons were usually unresponsive to the sensory stimuli used, many non-serotonergic neurons responded to these stimuli. Several non-serotonergic neurons showed a phasic relationship with splenius muscle activity during auditory stimulation. One serotonergic neuron showed a tonic relationship with digastric muscle activity during drinking behavior. A few non-serotonergic neurons exhibited a tonic relationship with digastric and/or masseter muscle activity during this behavior. Many non-serotonergic neurons exhibited a phasic relationship with these muscle activities, also during this behavior. These results suggest that the serotonergic neurons in the NRP and NRO constitute a relatively homogeneous population from a functional point of view, while the non-serotonergic neurons form groups with considerable functional specificity. The data support the idea that the NRP and NRO are implicated in the control of somatic motor output.

Breathing pattern, thoracoabdominal motion and muscular activity during three breathing exercises

The objective of the present study was to evaluate breathing pattern, thoracoabdominal motion and muscular activity during three breathing exercises: diaphragmatic breathing (DB), flow-oriented (Triflo II) incentive spirometry and volume-oriented (Voldyne) incentive spirometry. Seventeen healthy subjects (12 females, 5 males) aged 23 ± 5 years (mean ± SD) were studied. Calibrated respiratory inductive plethysmography was used to measure the following variables during rest (baseline) and breathing exercises: tidal volume (Vt), respiratory frequency (f), rib cage contribution to Vt (RC/Vt), inspiratory duty cycle (Ti/Ttot), and phase angle (PhAng). Sternocleidomastoid muscle activity was assessed by surface electromyography. Statistical analysis was performed by ANOVA and Tukey or Friedman and Wilcoxon tests, with the level of significance set at P < 0.05. Comparisons between baseline and breathing exercise periods showed a significant increase of Vt and PhAng during all exercises, a significant decrease of f during DB and Voldyne, a significant increase of Ti/Ttot during Voldyne, and no significant difference in RC/Vt. Comparisons among exercises revealed higher f and sternocleidomastoid activity during Triflo II (P < 0.05) with respect to DB and Voldyne, without a significant difference in Vt, Ti/Ttot, PhAng, or RC/Vt. Exercises changed the breathing pattern and increased PhAng, a variable of thoracoabdominal asynchrony, compared to baseline. The only difference between DB and Voldyne was a significant increase of Ti/Ttot compared to baseline. Triflo II was associated with higher f values and electromyographic activity of the sternocleidomastoid. In conclusion, DB and Voldyne showed similar results while Triflo II showed disadvantages compared to the other breathing exercises.

A simple test of muscle coactivation estimation using electromyography

In numerous motor tasks, muscles around a joint act coactively to generate opposite torques. A variety of indexes based on electromyography signals have been presented in the literature to quantify muscle coactivation. However, it is not known how to estimate it reliably using such indexes. The goal of this study was to test the reliability of the estimation of muscle coactivation using electromyography. Isometric coactivation was obtained at various muscle activation levels. For this task, any coactivation measurement/index should present the maximal score (100% of coactivation). Two coactivation indexes were applied. In the first, the antagonistic muscle activity (the lower electromyographic signal between two muscles that generate opposite joint torques) is divided by the mean between the agonistic and antagonistic muscle activations. In the second, the ratio between antagonistic and agonistic muscle activation is calculated. Moreover, we computed these indexes considering different electromyographic amplitude normalization procedures. It was found that the first algorithm, with all signals normalized by their respective maximal voluntary coactivation, generates the index closest to the true value (100%), reaching 92 ± 6%. In contrast, the coactivation index value was 82 ± 12% when the second algorithm was applied and the electromyographic signal was not normalized (P < 0.04). The new finding of the present study is that muscle coactivation is more reliably estimated if the EMG signals are normalized by their respective maximal voluntary contraction obtained during maximal coactivation prior to dividing the antagonistic muscle activity by the mean between the agonistic and antagonistic muscle activations.

Activation of neural cholecystokinin-1 receptors induces relaxation of the isolated rat duodenum which is reduced by nitric oxide synthase inhibitors

Cholecystokinin (CCK) influences gastrointestinal motility, by acting on central and peripheral receptors. The aim of the present study was to determine whether CCK has any effect on isolated duodenum longitudinal muscle activity and to characterize the mechanisms involved. Isolated segments of the rat proximal duodenum were mounted for the recording of isometric contractions of longitudinal muscle in the presence of atropine and guanethidine. CCK-8S (EC50: 39; 95% CI: 4.1-152 nM) and cerulein (EC50: 58; 95% CI: 18-281 nM) induced a concentration-dependent and tetrodotoxin-sensitive relaxation. Nomeganitro-L-arginine (L-NOARG) reduced CCK-8S- and cerulein-induced relaxation (IC50: 5.2; 95% CI: 2.5-18 µM) in a concentration-dependent manner. The magnitude of 300 nM CCK-8S-induced relaxation was reduced by 100 µM L-NOARG from 73 ± 5.1 to 19 ± 3.5% in an L-arginine but not D-arginine preventable manner. The CCK-1 receptor antagonists proglumide, lorglumide and devazepide, but not the CCK-2 receptor antagonist L-365,260, antagonized CCK-8S-induced relaxation in a concentration-dependent manner. These findings suggest that CCK-8S and cerulein activate intrinsic nitrergic nerves acting on CCK-1 receptors in order to cause relaxation of the rat duodenum longitudinal muscle.

Use of the caffeine-halothane contracture test for the diagnosis of malignant hyperthermia in Brazil

Malignant hyperthermia (MH) is a pharmacogenetic disease triggered by volatile anesthetics and succinylcholine. Deaths due to MH have been reported in Brazil. The first Malignant Hyperthermia Diagnostic and Research Center in Latin America was inaugurated in 1993 at the Federal University of Rio de Janeiro, Brazil. The center followed the diagnostic protocols of the North America MH Group, in which the contractures of biopsies from the vastus lateralis muscle are analyzed after exposure to caffeine and halothane (CHCT). CHCT was performed in individuals who survived, their relatives and those with signs/symptoms somewhat related to MH susceptibility (MHS). Here, we report data from 194 patients collected over 16 years. The Southeast (N = 110) and South (N = 71) represented the majority of patients. Median age was 25 (4-70) years, with similar numbers of males (104) and females (90). MHS was found in 90 patients and 104 patients were normal. Abnormal responses to both caffeine and halothane were observed in 59 patients and to caffeine or halothane in 20 and 11 patients, respectively. The contracture of biopsies from MHS exposed to caffeine and halothane was 1.027 ± 0.075 g (N = 285) and 4.021 ± 0.255 g (N = 226), respectively. MHS was found in patients with either low or high blood creatine kinase and also, with a low score on the clinical grading scale. Thus, these parameters cannot be used with certainty to predict MHS. We conclude that the CHCT protocol described by the North America MH Group contributed to identification of MHS in suspected individuals at an MH center in Brazil with 100% sensitivity and 65.7% specificity.

Characterization of angiotensin-converting enzymes 1 and 2 in the soleus and plantaris muscles of rats

Angiotensin-converting enzymes 1 (ACE1) and 2 (ACE2) are key enzymes of the renin-angiotensin system, which act antagonistically to regulate the levels of angiotensin II (Ang II) and Ang-(1-7). Considerable data show that ACE1 acts on normal skeletal muscle functions and architecture. However, little is known about ACE1 levels in muscles with different fiber compositions. Furthermore, ACE2 levels in skeletal muscle are not known. Therefore, the purpose of this study was to characterize protein expression and ACE1 and ACE2 activities in the soleus and plantaris muscles. Eight-week-old female Wistar rats (N = 8) were killed by decapitation and the muscle tissues harvested for biochemical and molecular analyses. ACE1 and ACE2 activities were investigated by a fluorometric method using Abz-FRK(Dnp)P-OH and Mca-YVADAPK(Dnp)-OH fluorogenic substrates, respectively. ACE1 and ACE2 protein expression was analyzed by Western blot. ACE2 was expressed in the skeletal muscle of rats. There was no difference between the soleus (type I) and plantaris (type II) muscles in terms of ACE2 activity (17.35 ± 1.7 vs 15.09 ± 0.8 uF·min-1·mg-1, respectively) and protein expression. ACE1 activity was higher in the plantaris muscle than in the soleus (71.5 ± 3.9 vs 57.9 ± 1.1 uF·min-1·mg-1, respectively). Moreover, a comparative dose-response curve of protein expression was established in the soleus and plantaris muscles, which indicated higher ACE1 levels in the plantaris muscle. The present findings showed similar ACE2 levels in the soleus and plantaris muscles that might result in a similar Ang II response; however, lower ACE1 levels could attenuate Ang II production and reduce bradykinin degradation in the soleus muscle compared to the plantaris. These effects should enhance the aerobic capacity necessary for oxidative muscle activity.

Effect of Propafenone on the Contractile Activity of Latissimus Dorsi Muscle Isolated in an Organ Chamber: Experimental Study in Rats

OBJECTIVE: To study the effect of propafenone on the contractile function of latissimus dorsi muscle isolated from rats in an organ chamber. METHODS: We studied 20 latissimus dorsi muscles of Wistar rats and divided them into 2 groups: group I (n=10), or control group - we studied the feasibility of muscle contractility; group II (n=10), in which the contralateral muscles were grouped - we analyzed the effect of propafenone on muscle contractility. After building a muscle ring, 8 periods of sequential 2-minute baths were performed, with intervals of preprogrammed electrical stimulation using a pacemaker of 50 stimuli/min. In group II, propafenone, at the concentration of 9.8 µg/mL, was added to the bath in period 2 and withdrawn in period 4. RESULTS: In group I, no significant depression in muscle contraction occurred up to period 5 (p>0.05). In group II, a significant depression occurred in all periods, except between the last 2 periods (p<0.05). Comparing groups I and II only in period 1, which was a standard period for both groups, we found no significant difference (p>0.05). CONCLUSION: Propafenone had a depressing effect on the contractile function of latissimus dorsi muscle isolated from rats and studied in an organ chamber.

Standardization of metachromatic staining method of myofibrillar ATPase activity of myosin to skeletal striated muscle of mules and donkeys

This study aims at standardizing the pre-incubation and incubation pH and temperature used in the metachromatic staining method of myofibrillar ATPase activity of myosin (mATPase) used for asses and mules. Twenty four donkeys and 10 mules, seven females and three males, were used in the study. From each animal, fragments from the Gluteus medius muscle were collected and percutaneous muscle biopsy was performed using a 6.0-mm Bergström-type needle. In addition to the metachromatic staining method of mATPase, the technique of nicotinamide adenine dinucleotide tetrazolium reductase (NADH-TR) was also performed to confirm the histochemical data. The histochemical result of mATPase for acidic pre-incubation (pH=4.50) and alkaline incubation (pH=10.50), at a temperature of 37ºC, yielded the best differentiation of fibers stained with toluidine blue. Muscle fibers were identified according to the following colors: type I (oxidative, light blue), type IIA (oxidative-glycolytic, intermediate blue) and type IIX (glycolytic, dark blue). There are no reports in the literature regarding the characterization and distribution of different types of muscle fibers used by donkeys and mules when performing traction work, cargo transportation, endurance sports (horseback riding) and marching competitions. Therefore, this study is the first report on the standardization of the mATPase technique for donkeys and mules.

Muscle sympathetic nerve activity and hemodynamic alterations in middle-aged obese women

To study the relationship between the sympathetic nerve activity and hemodynamic alterations in obesity, we simultaneously measured muscle sympathetic nerve activity (MSNA), blood pressure, and forearm blood flow (FBF) in obese and lean individuals. Fifteen normotensive obese women (BMI = 32.5 ± 0.5 kg/m²) and 11 age-matched normotensive lean women (BMI = 22.7 ± 1.0 kg/m²) were studied. MSNA was evaluated directly from the peroneal nerve by microneurography, FBF was measured by venous occlusion plethysmography, and blood pressure was measured noninvasively by an autonomic blood pressure cuff. MSNA was significantly increased in obese women when compared with lean control women. Forearm vascular resistance and blood pressure were significantly higher in obese women than in lean women. FBF was significantly lower in obese women. BMI was directly and significantly correlated with MSNA, blood pressure, and forearm vascular resistance levels, but inversely and significantly correlated with FBF levels. Obesity increases sympathetic nerve activity and muscle vascular resistance, and reduces muscle blood flow. These alterations, taken together, may explain the higher blood pressure levels in obese women when compared with lean age-matched women.

The influence of fasting/refeeding on the lipoprotein lipase activity of adipose tissue and muscle

Lipoprotein lipase activity in adipose tissue and muscle is modulated by changes in the pattern of food intake. We have measured total lipoprotein lipase activity in adipose tissue and muscle of male Wistar rats (N = 6-10), weighing 200-250 g (~12 weeks), during the refeeding/fasting state following 24 h of fasting. Lipoprotein lipase activity in tissue homogenates was evaluated using a [³H]-triolein-containing substrate, and released [³H]-free fatty acids were extracted and quantified by liquid scintillation. Adipose tissue lipoprotein lipase activity did not completely recover within 2 h of refeeding (60% of refed ad libitum values). Cardiac lipoprotein lipase activity remained increased even 2 h after refeeding (100% of refed ad libitum values), whereas no significant changes were observed in the soleus and diaphragm muscles. Adipose tissue lipoprotein lipase activities were consistently higher than the highest skeletal muscle or heart values. It is therefore likely that adipose tissue, rather than muscle makes the major contribution to triacylglycerol clearance. There was concomitant relatively high lipoprotein lipase activity in both adipose tissue and cardiac muscle during the first few hours of refeeding, therefore cardiac muscle may contribute significantly to triacylglycerol clearance during this period. The results suggest that during fasting, increased lipoprotein lipase activity provides a complementary source of free fatty acids from circulating triacylglycerol, allowing the heart to maintain its continuous, high-energy expenditure.

Effects of lipopolysaccharide on low- and high-density cultured rabbit vascular smooth muscle cells: differential modulation of nitric oxide release, ERK1/ERK2 MAP kinase activity, protein tyrosine phosphatase activity, and DNA synthesis

Previous studies have shown that exogenously generated nitric oxide (NO) inhibits smooth muscle cell proliferation. In the present study, we stimulated rabbit vascular smooth muscle cells (RVSMC) with E. coli lipopolysaccharide (LPS), a known inducer of NO synthase transcription, and established a connection between endogenous NO, phosphorylation/dephosphorylation-mediated signaling pathways, and DNA synthesis. Non-confluent RVSMC were cultured with 0, 5, 10, or 100 ng/ml of the endotoxin. NO release was increased by 86.6% (maximum effect) in low-density cell cultures stimulated with 10 ng/ml LPS as compared to non-stimulated controls. Conversely, LPS (5 to 100 ng/ml) did not lead to enhanced NO production in multilayered (high density) RVSMC. DNA synthesis measured by thymidine incorporation showed that LPS was mitogenic only to non-confluent RVSMC; furthermore, the effect was prevented statistically by aminoguanidine (AG), a potent inhibitor of the inducible NO synthase, and oxyhemoglobin, an NO scavenger. Finally, there was a cell density-dependent LPS effect on protein tyrosine phosphatase (PTP) and ERK1/ERK2 mitogen-activated protein (MAP) kinase activities. Short-term transient stimulation of ERK1/ERK2 MAP kinases was maximal at 12 min in non-confluent RVSMC and was prevented by preincubation with AG, whereas PTP activities were inhibited in these cells after 24-h LPS stimulation. Conversely, no significant LPS-mediated changes in kinase or phosphatase activities were observed in high-density cells. LPS-induced NO generation by RVSMC may switch on a cell density-dependent proliferative signaling cascade, which involves the participation of PTP and the ERK1/ERK2 MAP kinases.

Eucalyptol, an essential oil, reduces contractile activity in rat cardiac muscle

Eucalyptol is an essential oil that relaxes bronchial and vascular smooth muscle although its direct actions on isolated myocardium have not been reported. We investigated a putative negative inotropic effect of the oil on left ventricular papillary muscles from male Wistar rats weighing 250 to 300 g, as well as its effects on isometric force, rate of force development, time parameters, post-rest potentiation, positive inotropic interventions produced by Ca2+ and isoproterenol, and on tetanic tension. The effects of 0.3 mM eucalyptol on myosin ATPase activity were also investigated. Eucalyptol (0.003 to 0.3 mM) reduced isometric tension, the rate of force development and time parameters. The oil reduced the force developed by steady-state contractions (50% at 0.3 mM) but did not alter sarcoplasmic reticulum function or post-rest contractions and produced a progressive increase in relative potentiation. Increased extracellular Ca2+ concentration (0.62 to 5 mM) and isoproterenol (20 nM) administration counteracted the negative inotropic effects of the oil. The activity of the contractile machinery evaluated by tetanic force development was reduced by 30 to 50% but myosin ATPase activity was not affected by eucalyptol (0.3 mM), supporting the idea of a reduction of sarcolemmal Ca2+ influx. The present results suggest that eucalyptol depresses force development, probably acting as a calcium channel blocker.

Effects of high-intensity intermittent training on carnitine palmitoyl transferase activity in the gastrocnemius muscle of rats

We examined the capacity of high-intensity intermittent training (HI-IT) to facilitate the delivery of lipids to enzymes responsible for oxidation, a task performed by the carnitine palmitoyl transferase (CPT) system in the rat gastrocnemius muscle. Male adult Wistar rats (160-250 g) were randomly distributed into 3 groups: sedentary (Sed, N = 5), HI-IT (N = 10), and moderate-intensity continuous training (MI-CT, N = 10). The trained groups were exercised for 8 weeks with a 10% (HI-IT) and a 5% (MI-CT) overload. The HI-IT group presented 11.8% decreased weight gain compared to the Sed group. The maximal activities of CPT-I, CPT-II, and citrate synthase were all increased in the HI-IT group compared to the Sed group (P < 0.01), as also was gene expression, measured by RT-PCR, of fatty acid binding protein (FABP; P < 0.01) and lipoprotein lipase (LPL; P < 0.05). Lactate dehydrogenase also presented a higher maximal activity (nmol·min-1·mg protein-1) in HI-IT (around 83%). We suggest that 8 weeks of HI-IT enhance mitochondrial lipid transport capacity thus facilitating the oxidation process in the gastrocnemius muscle. This adaptation may also be associated with the decrease in weight gain observed in the animals and was concomitant to a higher gene expression of both FABP and LPL in HI-IT, suggesting that intermittent exercise is a "time-efficient" strategy inducing metabolic adaptation.

Microbicidal effect of medicinal plant extracts (Psidium guajava Linn. and Carica papaya Linn.) upon bacteria isolated from fish muscle and known to induce diarrhea in children

Out of the twenty-four samples of shrimp and fish muscle used for this study, twelve were collected near a large marine sewer for waste disposal, 3 km off the coast of Fortaleza (Brazil) and used for the isolation of E. coli. Other twelve were collected at the Mucuripe fresh fish market (Fortaleza, Brazil) and used for the isolation of Staphylococcus aureus. Ethanol, water and acetone-diluted extracts of guava and papaya leaf sprouts were tested on the bacteria in order to verify their microbicidal potential. The E. coli strains used in the trials were rated LT positive. The papaya leaf extracts (Carica papaya Linn) showed no microbicidal activity while the guava sprout extracts (Psidium guajava Linn) displayed halos exceeding 13 mm for both species, an effect considered to be inhibitory by the method employed. Guava sprout extracts by 50% diluted ethanol most effectively inhibited E. coli (EPEC), while those in 50% acetone were less effective. It may be concluded that guava sprout extracts constitute a feasible treatment option for diarrhea caused by E. coli or by S. aureus-produced toxins, due to their quick curative action, easy availability in tropical countries and low cost to the consumer.