Abstract
We have shown that vasopressinergic projections to dorsal brain stem are activated during exercise and facilitate exercise tachycardia in both trained (T) and sedentary (S) rats (Dufloth DL, Morris M, and Michelini LC. Am J Physiol Regulatory Integrative Comp Physiol 273: R1271–R1282, 1997). In the present study, we investigated whether oxytocinergic projections to the nucleus of the solitary tract (NTS)-dorsal motor nucleus of the vagus (DMV) complex (NTS/DMV) are involved in the differential heart rate (HR) response to exercise in T and S rats. Arterial pressure (AP) and HR responses to dynamic exercise (0.4–1.4 km/h) were compared in S and T pretreated with vehicle (saline), oxytocin (OT; 20 pmol/200 nl) or OT-receptor antagonist (OTant; 20 pmol/200 nl) into the NTS/DMV. OT content in specific brain regions and plasma were measured in separate S and T groups at rest and immediately after exercise. Exercise increased OT content in dorsal (4.5-fold) and ventral brain stem (2.7-fold) and spinal cord (3.4-fold) only in T rats. No significant changes were observed in neurosecretory regions or medial eminence and posterior pituitary, but plasma levels of T rats were reduced immediately after exercise. Blockade of NTS/DMV OT receptors did not change basal mean AP (MAP) and HR or the MAP response to exercise. However, OTant potentiated exercise-induced tachycardia (average increase of 26%) only in the T group. Pretreatment with exogenous OT in the NTS/DMV blunted the tachycardic response both in S and T rats without changing the MAP response. Administration of OT-receptor antagonist or OT into the fourth cerebral ventricle had no effect on the cardiovascular response to dynamic exercise. Taken together, the results suggest that oxytocinergic projections to the NTS/DMV are stimulated when T rats exercise and that OT released at this level acts on OT receptors to restrain exercise-induced tachycardia.
- oxytocin receptors
- nucleus of the solitary tract
- dorsal motor nucleus of the vagus
- fourth ventricle
- blood pressure
- trained rats
the cardiovascular response to exercise is characterized by a prompt moderate rise in blood pressure accompanied by marked tachycardia. Reflex bradycardia is absent, although experimental evidence demonstrates that baroreceptors are active during exercise (13, 40). When baroreceptors were stimulated in rats running on a treadmill, there was a significant reflex bradycardia (13). The sensitivity of the bradycardic response after an acute bout of exercise was similar to that seen during rest (40). It has been proposed (5, 35) that reflex sensitivity is maintained during exercise by resetting of the operating point of the arterial baroreflex to higher pressures.
The nucleus of the solitary tract (NTS, first synaptic relay of baroreceptors) and the dorsal motor nucleus of the vagus (DMV, the origin of parasympathetic preganglionic neurons) are important brain stem areas involved in the reflex control of cardiovascular function. Several peptides have been identified in terminals present in the NTS/DMV complex (44), some of hypothalamic origin (27, 38, 42). Our initial studies focused on the role of brain stem vasopressin in the modulation of exercise-induced cardiovascular changes. The direct administration of vasopressin into the NTS of conscious rats shifted the operating point of baroreceptor reflex control of heart toward higher heart rate (HR) values without altering reflex sensitivity (20). In addition, we showed that vasopressinergic projections from paraventricular nucleus (PVN) to dorsal brain stem (DBS) are activated during exercise as demonstrated by the increase in vasopressin content and by the blunting of exercise tachycardia after vasopressin antagonist administration into the NTS (7, 19). We postulated that vasopressinergic projections to the NTS act as one of the links between the “central command” for the motor activity and the circulatory control driven by brain stem feedback mechanisms (19, 22). However, endogenous vasopressin does not account for all of the effects, because pretreatment with a receptor antagonist blunted, but did not block, exercise-induced tachycardia (7). Furthermore, the difference in HR response observed in trained animals (smaller tachycardia) was not correlated with the change in central vasopressin content (7). In humans, training alters exercise tachycardia with lower HR responses to submaximal exercise observed in trained versus sedentary individuals (4, 39). This adaptive effect of training is accompanied by changes in vagal and sympathetic tone to the heart (25, 26, 39, 41) and perhaps alterations in central peptidergic systems.
Oxytocin (OT), a peptide similar to vasopressin in terms of structure and synthesis sites, is known primarily for its role in milk ejection, parturition, and behavior (30). However, in brain stem regions (NTS and DMV), the concentrations of OT and its receptor are much higher than vasopressin (6, 10, 27, 38, 43). OT also has cardiovascular effects, such as modulation of stress-induced tachycardia (24) and baroreflex sensitivity (28, 36). In a preliminary report (23), we observed that OT administered into the NTS/DMV complex potentiated the reflex bradycardia and blunted the tachycardic response to acute exercise, responses that were opposite to that seen for vasopressin. Together, these observations suggest that central OT is a potential candidate as a modulator of HR control during exercise.
The objectives of the present study were 1) to determine the effect of dynamic exercise on OT content in specific brain regions and plasma; 2) to investigate the effect of OT-receptor blockade in both NTS/DMV and fourth cerebral ventricle on blood pressure and HR responses to treadmill exercise; 3) to determine the effects of injection of OT into the same areas on the cardiovascular responses to exercise; and 4) to compare those responses in sedentary (S) and trained (T) rats. The experimental protocol involves a technique developed in our laboratory (20) in which drugs are microinjected into the DBS of conscious, freely moving rats. This allows for the evaluation of central OT actions on cardiovascular responses during treadmill exercise.
MATERIALS AND METHODS
Animals/Training Protocol
Male Wistar-Kyoto (WKY) rats, weighing 170–200 g at the beginning of the experiments, were housed four per cage and kept on a 12:12-h light-dark cycle with standard laboratory chow and water ad libitum. Body weight was measured weekly. All surgical procedures and protocols used were in accordance with Guidelines for Ethical Care of Experimental Animals and were approved by the Institutional Animal Care and Use Committee.
Rats were initially preselected for their ability to walk on a treadmill (Inbramed, KT-300, 4–5 sessions at 0.3–0.5 km/h, 0% grade, and 10 min/day) and then randomly assigned to T or S groups. The training protocol was based on one first described by Negrao et al. (26), and modified in our laboratory (7). The exercise is progressive with increased exercise intensity, determined by a combination of velocity (0% grade) and time. Exercise was performed 5 days/wk for 13 wk. It began with 0.3 km/h, 10–15 min/day, reaching 0.5 km/h and 30 min/day in week 1. The training intensity was gradually increased to a duration of 60 min/day (by week 3) with a speed of 1.0 km/h (by week 7), a schedule which was continued until the end of the experiment. The training protocol resulted in 50–60% of maximal oxygen uptake (V˙o 2 max), as determined in some rats before the 13-wk period and again by the middle of the training protocol. The protocol used for O2consumption testing was that employed by Silva et al. (40). Briefly, it consisted of the measurement ofV˙o 2 max by means of expired gas analysis during both rest and a progressive exercise test on a treadmill (increments of 0.3 km/h, every 4 min until exhaustion). Expired gas samples were collected during the last 30 s in each load; oxygen and carbon dioxide concentrations in the samples were analyzed with Scholander microtechnique for gas analysis. It should be noted that the metabolic capacity, as indicated byV˙o 2 max measured in four rats of each group, was increased by 47% in T groups compared with S groups (average of 89 ± 5 ml O2 ⋅ kg−1 ⋅ min−1). Rats allocated to the S protocol were handled every day and submitted once a week to a short period of mild exercise (5–10 min, 0.4–0.8 km/h, 0% grade).
Measurement of Plasma and Tissue OT
To determine the effect of exercise on brain and plasma OT levels, one-half of T and one-half of S rats were killed by decapitation when resting in their home cage, whereas the remaining rats were killed immediately after performing graded exercise tests on the treadmill (0.4, 0.8, and 1.1 km/h for S and 0.4, 0.8, 1.1, and 1.4 km/h for T rats, 2 min each load). These exercise protocols resulted in maximal HR responses. Trunk blood was collected in heparinized tubes on ice, centrifuged to separate the plasma, and stored at −80°C. The brain and posterior pituitary (PP) were rapidly removed, placed in dry ice-cooled isopentane, frozen in liquid nitrogen, and stored at −80°C. PVN and supraoptic (SON) nuclei, median eminence (ME), DBS and ventral brain stem (VBS), and spinal cord (SC) regions were microdissected from frozen brain sections. The tissue samples were weighed and sonicated in HCl (0.1 N), and peptide content was measured. Plasma samples were extracted with acetone precipitation and petroleum ether extraction. Plasma and tissue extracts were measured for OT by a sensitive and specific radioimmunoassay (32). The rabbit antiserum is specific for the amidated peptide, with negligible cross-reactivity with vasopressin or other related peptides. The peptide standard and [125I]OT were purchased from commercial sources (Bachem, Torrance, CA and DuPont, Boston, MA, respectively).
Cardiovascular Measurements
Separate groups of T and S rats were used for the cardiovascular studies. Guide cannulas were inserted into the DBS according to a technique developed in our laboratory (20). Briefly, the rats were anesthetized with pentobarbital sodium (40 mg/kg ip) and placed in a stereotaxic apparatus (Kopf, Tujunga, CA), with their heads in a horizontal position. The skull was exposed, a screw was inserted into the parietal bone, and a small window was opened caudally to the lambda to allow the introduction of a stainless steel guide cannula (17 mm length, 0.6 mm OD) at an angle of 24°. The stereotaxic coordinates for NTS/DMV placement were as follows: 1 mm caudal to interaural line, 0.4 mm lateral (right or left) to the midline, and 8.9 mm below the skull surface. For the fourth ventricle cannulation, the rostrocaudal and lateral coordinates were the same, but the guide cannula was introduced 8.8 mm. It is important to note that the tip of the guide cannula did not reach the NTS/DMV area, but lay in the ventral cerebellum, avoiding extensive tissue damage to the areas of interest. The cannula was fixed in place with fast polymerizing methacrylate cement, and an occluder closed the guide cannula. The rats were given 60,000 units of penicillin (Pentabiotico Veterinario, Fontoura Wyeth, Sao Paulo, SP, Brazil). After a recovery period of 2–3 days, the training program was restarted. Six to eight days after brain stem cannulation and 1 day before the acute running tests, a femoral arterial catheter (3 cm of PVC tube OD 0.61 × ID 0.28 mm, connected to 12 cm of PVC tube OD 1.50 × ID 0.50 mm, Critchley, Silverwater, Australia) was inserted under ether anesthesia. The catheter was tunneled subcutaneously to the back of the neck, where it was fixed with a suture. It was flushed with heparinized saline 6–8 h later.
The experimental design consisted of testing the cardiovascular responses to exercise in two specific situations: 1) after administration of OT antagonist [deamino-Cys1,d-Tyr (Et)2,Thr4,Orn8]-oxytocin (OTant, Bachem) at a dose of 20 pmol/200 nl into the NTS/DMV or fourth ventricle and 2) after administration of OT at a dose of 20 pmol/200 nl into the same areas. In both protocols, vehicle (Veh; 0.9% NaCl, 200 nl) was administered for the control. Because of the long-lasting effects of centrally administered OT (29), different groups of rats were used for OTant and OT administration into the NTS/DMV.
During the experimental sessions, arterial pressure (systolic, diastolic, and mean) and HR (computed from the arterial pressure pulses) were recorded continuously (P23XL transducer, connected to RS3400 recorder, Gould, Cleveland, OH). At the beginning of the experimental session, the rat was placed on the treadmill and the occluder was removed from the guide cannula. Twenty to thirty minutes were allowed for stabilization of the cardiovascular parameters. Microinjections were made by introduction of a 33-gauge needle (18 mm length) connected by PE-10 tubing to a microliter syringe (701-N, Hamilton, Reno, NV) into the guide cannula. Microinjections lasted 15–20 s and the needle was removed 5–10 s later. The rats rested for another 20–30 min, when baseline values of mean arterial pressure (MAP) and HR were obtained. The acute exercise tests consisted of graded exercise intensities (labeled mild, moderate, or heavy when maximal HR responses were obtained), maintained for 2 min each: 0.4, 0.8, and 1.1 km/h for S and 0.4, 0.8, 1.1, and 1.4 km/h for T rats. The exercise test was followed by a 40-min recovery period. For each animal, the protocol was repeated twice: for Veh and OT or OTant treatments. At the end of the experimental session, 200 nl of Evans blue dye were microinjected into the brain stem. The rat was then anesthetized with ether and perfused transcardially with 30–40 ml of saline followed by 10% buffered Formalin. The brain was removed and stored in 10% Formalin plus 10% sucrose. The exact location of the injection site and its extension were assessed a posteriori by histological examination of 40-μm serial coronal sections obtained with a cryostat (Sartorius-Werke, Gottingen, Germany).
Data Analysis and Statistics
The data are presented as means ± SE. Functional data are reported as absolute resting values and changes of MAP and HR from rest values for S and T rats during exercise and recovery. A three-way analysis of variance (treatment × exercise intensity × time) with repeated measures on the third factor (BMDP, Statistical Software) was used to compare HR and MAP changes in S and T groups (time-course effects). Maintained or steady-state HR and MAP responses, determined during the last minute in each exercise load in S and T groups, were also compared by the three-way analysis of variance using group × treatment × exercise intensity as the factor design. Differences of plasma and central nervous system OT levels between groups (S and T) and conditions (rest and exercise) were evaluated by two-way analysis of variance. Tukey's test was used as a post hoc test. The level of significance was P < 0.05.
RESULTS
Brain and Plasma OT: Effect of Training and Exercise
Measurements of regional brain OT (nuclear and terminal projection areas) and plasma OT content showed that there were no significant differences between S and T rats under resting conditions (Tukey's test, Table 1). A close observation on OT levels revealed that after training, resting PVN content was increased by 72% (group effect, P < 0.05) and that plasma level was increased by 37% (group × condition interaction, P< 0.05). In the T group, immediately after dynamic exercise, there was a marked increment in the DBS OT content (+1.7 ± 0.5 pg/mg tissue, a 4.5-fold increase) and significant increases in VBS and SC (+0.8 ± 0.2 and +0.6 ± 0.1 pg/mg tissue or 2.7- and 3.4-fold increases, respectively). These changes were specific for the T group, because in the S rats, DBS, VBS, and SC postexercise OT levels were not different from the values observed at rest (Table 1). Exercise did not change OT content in the biosynthetic areas (PVN, SON) or in the areas corresponding to magnocellular pathways (PP, ME). In the T group, however, plasma OT was reduced immediately after the exercise on the treadmill (Table 1).
Brain and plasma oxytocin content at rest and immediately after dynamic exercise in sedentary and trained rats
Cardiovascular Responses to Exercise: Effect of Injection of OTant and OT
Resting conditions.
Control values of MAP and HR at rest in S and T chronically cannulated groups pretreated with Veh are shown in Table2. Pre-Veh control measurements of MAP and HR were also included, showing that Veh and volume (200 nl) injections into the NTS/DMV were without effect. Pretreatment with OTant (20 pmol/200 nl, groups 1 and 2) or OT (20 pmol/200 nl, groups 3 and 4) into the NTS/DMV did not change significantly the basal values of MAP and HR of the conscious rats at rest.
Resting values of MAP and HR in sedentary and trained rats before and after Veh pretreatment
Cardiovascular response to exercise in S and T rats.
In both groups, the control (Veh) MAP response to treadmill exercise was characterized by an initial peak increase followed by a smaller but maintained MAP increase (10- to 20-mmHg range, Fig.1). The HR response also showed an initial large increment followed by a gradual and maintained increase, with the magnitude related to the exercise load. The maximal HR response was similar for S and T groups (140 ± 12 and 149 ± 8 beats/min over baseline, respectively) but was attained at different exercise loads (1.1 and 1.4 km/h for S and T rats, respectively).
Heart rate (HR; top) and mean arterial pressure (MAP;bottom) responses during different exercise intensities and recovery in sedentary (S; left, n = 8) and trained rats (T; right, n = 8) pretreated with vehicle (Veh) and oxytocin (OT) antagonist (OTant) into nucleus of the solitary tract (NTS)-dorsal motor nucleus of the vagus (DMV) complex (NTS/DMV). * Significance (P < 0.05) vs. Veh.
Exercise after pretreatment with OTant or OT.
Injection of OTant in the NTS/DMV did not change the MAP response to exercise in either group (Fig. 1), neither did it change the HR response of S rats (Fig. 1, left). However, OT-receptor blockade restricted to the NTS/DMV area significantly potentiated the exercise tachycardia of T rats (Fig. 1, right). In the T group, the maximal HR response at 1.4 km/h was +187 ± 6 beats/min, an increment of 26% over the maximal value observed after Veh treatment (+149 ± 8 beats/min).
The HR responses at steady-state exercise during the different exercise intensities in both S and T groups pretreated with Veh and OTant or Veh and OT into the NTS/DMV are compared in Fig.2. In the T group, pretreatment with OTant increased exercise tachycardia (Fig. 2 B): increments ranged from +15% to +30%, with significant changes being observed at 0.8, 1.1, and 1.4 km/h. There was no significant HR effect when S rats were pretreated with OTant (Fig. 2 A). On the other hand, OT given into the NTS/DMV caused significant blunting of exercise tachycardia in both groups: −37%, −23%, −19% for 0.4–1.1 km/h (Fig. 2 C) and −35%, −18%, −17%, −22% for 0.4–1.4 km/h (Fig. 2 D), S and T groups, respectively. OTantand OT treatments in both S and T groups did not cause any significant change in MAP response at steady-state exercise during different exercise intensities (data not shown).
HR responses at steady-state exercise during different workloads after Veh, OTant, and OT treatments into NTS/DMV of S (Aand C) and T (B and D) rats. Different groups of rats were used to test OTant (n = 8 in Aand B) and OT effects (n = 9 for C, n = 8 for D). All changes from respective baseline HR were significant; * significant difference from Veh treatment (P< 0.05).
To understand the effects of facilitation and/or inhibition of oxytocinergic synapses at NTS/DMV on exercise tachycardia and the changes caused by training, we compare in Fig.3 the differential effect of OTant and OT (after subtraction of the tachycardic response observed after Veh pretreatment). The effect of OT-receptor blockade on exercise tachycardia differs dramatically in T and S groups: marked improvement versus no change. This does not mean that OT has no effect in the NTS/DMV of S rats, because exogenous administration caused the same inhibitory effect in both groups (Fig. 3). The data indicate that in the S group, brain stem OT input is not activated during exercise, as supported by the lack of change in OT content immediately after the exercise test.
Comparison of oxytocinergic effects on exercise tachycardia at different workloads in S and T groups. Bars represent differential HR effect of both antagonist [OTant − Veh responses (A)] and agonist [OT − Veh responses (B)] pretreatments into NTS/DMV. All HR changes, except those after OTant treatment in S rats, were significant. * Difference between S and T groups (P < 0.05).
Fourth ventricular injections (Table 3) were used to test the specificity of the responses and to compare the brain stem versus the cerebrospinal fluid (CSF) effects (local neuronal-released OT versus OT released into the CSF). As described previously for the NTS/DMV groups, Veh administration into the CSF (200 nl) did not change resting values of MAP and HR. Also, MAP and HR responses after Veh pretreatment in the fourth cerebral ventricle were similar to those observed when Veh was given into the NTS/DMV. Exogenous OT (20 pmol/200 nl) or its endogenous blockade in the CSF caused no change in the exercise tachycardia or MAP response to exercise.
Resting values and MAP and HR response during dynamic graded exercise in sedentary and trained rats
Injection sites.
Microscopic examination of the brain stem in all groups of S and T rats revealed that in all rats included in this study, the microinjections were directed to the commissural NTS and adjacent DMV. There was a great coincidence of microinjected areas between S and T groups. The dye injected apparently followed the elongated structure of both NTS and DMV by spreading predominantly in the rostrocaudal direction. The average extension of dye-stained areas was 455 ± 51 μm in S and 448 ± 43 μm in T groups, and for both groups the mean point of injection was located between the obex and 200 μm rostral. In the rats in which the injections targeted the fourth ventricle, macroscopic examination immediately after removal of the brain revealed the presence of the dye in CSF. In some rats, the dye was also present in the dorsal region of the spinal cord. In these rats, histological studies showed no injection sites in the brain stem.
DISCUSSION
These findings provide evidence that OT, a peptide known for its role in maternal function, is also important in the regulation of the cardiovascular system. These data demonstrate that central OT modulates the HR response to exercise. Specifically, the study provides several new observations: 1) exercise produces a specific increase in the brain stem and spinal cord OT content in T rats; 2) blockade of OT receptors in the NTS/DMV does not change MAP response but potentiates the tachycardic response only in the T group;3) injection of OT into the NTS/DMV blunts the tachycardic response in both S and T groups without changing MAP response; and 4) OT injection into the fourth cerebral ventricle does not affect the cardiovascular responses to dynamic exercise.
It is well known that in normotensive individuals, exercise training increases maximal oxygen uptake that is accompanied by cardiovascular adaptations such as increase in myocardium contractility and cardiac output, increase in the capillary supply and flow to exercised muscles, and increased oxygen extraction by the working muscles (1, 4, 39). These cardiac and skeletal muscle changes occur without alterations in resting blood pressure. It is also well established that during exercise, trained individuals show maximal HR responses, which are similar to sedentary individuals. However, the maximal responses are attained at a larger exercise load with smaller tachycardic responses seen at submaximal exercise intensities (4, 31). The combined actions of parasympathetic withdrawal, an immediate response, and sympathetic activation, the main effect at steady-state exercise (11, 25, 31), are responsible for exercise-induced tachycardia. Training was shown to decrease the HR response by reducing intrinsic HR (26) and changing the autonomic control of the heart (11, 25, 39, 41).
The central mechanism(s) responsible for these exercise-induced changes has not been identified. On the basis of our data, we hypothesize that OT released into the DBS could play a role, acting to reduce tachycardia and the resulting energy expense without changing the blood pressure response. This effect appears to be specific for the trained situation. There is limited and sometimes controversial information on the role of OT in cardiovascular regulation. In the periphery, it acts as a natriuretic agent and a weak vasoconstrictor (30). In the central nervous system, it has multiple actions on HR and blood pressure regulation. For example, intraventricular OT administration has been shown to increase HR (28) or have no significant cardiovascular effects (9). When injected into the NTS, OT caused tachycardia and hypertension (18), whereas a bradycardia resulted on injection into the DMV (33). These effects appeared to be specific, because they were antagonized by central OT-receptor blockade (34). In addition, OT given intravenously or into the CSF has been shown to alter baroreceptor reflex control of the heart (28, 36). New studies in our laboratory showed prominent differences in autonomic control of the heart in mice lacking the ability to synthesize OT, so-called OT knockout (OTKO) (21). The OTKO mice showed an altered baroreflex function curve with increased sensitivity and reduced baseline HR. In the absence of OT gene, reflex bradycardia was significantly blunted during blood pressure increase, results compatible with the attenuated exercise-induced tachycardia after OT treatment. Likewise, central injection of OT antisense reduced OT peptide levels and attenuated the HR response to stress and substance P (16, 21).
Neuronal tracers and immunohistochemical studies showed that hypothalamic parvocellular PVN neurons project to the NTS and DMV in the DBS, to the ventrolateral medulla in the VBS, and to the intermediolateral cell column of the spinal cord, areas known to be involved in the autonomic control of the heart and vasculature (2, 27,38, 42, 44). Previous work demonstrated that the PVN is the primary source of OT in the brain stem and spinal cord via projections from parvocellular neurons (2, 27, 38, 42). PVN lesions, specifically of parvocellular neurons, produced a decrease in OT content in the brain stem and spinal cord, whereas SON lesions produced no changes (3, 12). It is interesting that exercise produced significant changes in brain stem and spinal cord OT content, an effect that occurred only in trained animals. There were significant increases in DBS (corresponding to the NTS/DMV), VBS [corresponding to ventrolateral medulla (VLM)], and SC (corresponding to OT projections to spinal cord) immediately after dynamic exercise tests. These regions could act together to modulate HR via effects on afferent input and vagal and sympathetic output. The question raised by these data is whether the changes in content are associated with alterations in peptide synthesis and secretion. However, there was specificity in the responses, because peptide content was increased only in the trained group and HR changes after endogenous OT blockade were seen only in trained rats. OT release during exercise was also regionally specific, because there were no changes in the neurosecretory regions (PVN or SON) or the terminal projections (PP or ME). Plasma levels were reduced in exercised, trained rats, suggesting some effect on magnocellular systems. Taken together, the results suggest that parvocellular oxytocinergic projections to the NTS/DMV are stimulated when trained rats exercise and that OT released at this level contributes to the smaller HR response seen in this group. Actually, parvocellular PVN neurons projecting to the DBS have been implicated in HR control, because PVN stimulation increases (34), whereas lesions attenuate responsiveness (3, 12). Support for brain stem-centered action is seen in the study of fourth ventricular injections. OT or OTant produced no effect on the cardiovascular responses to exercise, suggesting that CSF OT is not involved in this response. It should be noted that immediately after running on the treadmill, OT content was also increased in VBS and SC. The functional effects of OT in the VBS (corresponding to VLM) and spinal cord during exercise remain to be determined.
Interestingly, when OT was administered into the NTS/DMV, the exercise tachycardia was similarly blunted in both sedentary and trained groups. This was in contrast to the results with OTant treatment, in which effects were seen only in the T group. There is evidence for a high density of OT receptors and fibers in the NTS and DMV (2, 6, 10,27, 38, 42-44). The results support the idea that physiologically active OT receptors are present in the NTS/DMV of both S and T rats and suggest that the agonist/receptor interaction inhibits exercise-induced tachycardia. The difference observed between S and T groups after OT-receptor blockade may be due to an enhancement of OT release in trained animals. Our results do not allow the identification of mechanisms (pre- or postsynaptic, intracellular pathways) involved in this response. It is possible that training induces changes in OT receptors/intracellular signaling as well as secretion.
It is important to recognize that the OT antagonist used in the present study is selective for OT receptors, with a high in vivo anti-OT/anti-V1a selectivity (17). We have shown that vasopressin is also released into the NTS of rats during exercise and that vasopressin injection into this area potentiates the HR response to exercise (7, 19, 22). Previous studies of static muscle contraction induced by stimulation of the ventral root in anesthetized cats also showed increases in blood pressure and HR associated with Fos activity in both PVN oxytocinergic and vasopressinergic neurons (14). The OT and vasopressin released in the DBS during dynamic exercise seem to have specific and opposite effects on HR control. Similar “arrangements” in which two closely related substances have opposite effects have been described in other biological systems such as the renin-angiotensin (angiotensin II × angiotensin 1–7; 8, 37) and those of the endothelium-derived factors (EDRFs × EDCFs) (15). A balance between excitatory and inhibitory stimuli improves the efficiency of the system to better adjust the physiological response to the momentary requirements. Therefore, at the NTS level, OT and vasopressin act as neurotransmitters to modulate HR response to exercise by decreasing or increasing, respectively the tachycardia. It is apparent from our previous (7) and present experiments that the oxytocinergic input predominates after training.
In summary, a training paradigm using chronically cannulated and monitored rats has demonstrated the importance of central OT in the modulation of HR during exercise. A comparison of the cardiovascular effects of agonist and antagonist treatment as well as measurements of OT in brain and plasma provide evidence that central OT is inhibitory to exercise tachycardia. Although autonomic adjustments to exercise induced by central OT have not been studied yet, we observed that improvement of reflex bradycardia after OT administration in the NTS/DMV was mediated by the vagal tone to the heart (L. C. Michelini, unpublished observations). Therefore, it is possible that the circulatory adjustments to exercise in T rats are achieved in the presence of autonomic balance that favors the parasympathetic control of the heart. The results of the present experiments suggest that extensive treadmill training produces alterations in central oxytocinergic systems, which may mediate its cardioprotective effects.
Perspectives
By combining in resting and exercising sedentary and trained rats the measurement of OT content in discrete brain stem areas with the functional responses to exercise after pretreatment of the solitarii-vagal complex with OT or its antagonist, we are able to document for the first time a central oxytocinergic mechanism that restrains exercise-induced tachycardia in trained individuals. This study, which bridges the disciplines of exercise physiology and neural control of circulation with biochemical measurements of tissue peptide content, is unique and highlights the physiological significance of oxytocinergic synapses in the solitarii-vagal complex to modulate HR (and cardiac output) control during exercise. More importantly, it opens a new field for studying the modulation of autonomic function during different behaviors by suprabulbar projections (OT, other peptidergic and/or catecholaminergic projections) to areas in the brain stem involved in the primary reflex control of the circulation.
Acknowledgments
Supported by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP-96/0043–4; 98/03603–6), Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq-523337/95–7) and National Heart, Lung, and Blood Institute Grant HL-43178.
Footnotes
Address for reprint requests and other correspondence: L. C. Michelini, Dept. of Physiology and Biophysics, ICB, Univ. of Sao Paulo, Av. Prof. Lineu Prestes, 1524, 05508–900 Sao Paulo, SP Brazil (E-mail:Michelin{at}usp.br).
L. C. Michelini is a Research Fellow from CNPq.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. §1734 solely to indicate this fact.
- Copyright © 2000 the American Physiological Society
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