Intracerebroventricular insulin injection acutely normalizes the augmented exercise pressor reflex in male rats with type 2 diabetes mellitus.

The exercise pressor reflex (EPR) is exaggerated in type 2 diabetes mellitus (T2DM), but the underlying central nervous system aberrations have not been fully delineated. Stimulation of muscle afferents within working skeletal muscle activates the EPR, by sending information to neurons in the brainstem, where it is integrated and results in reflexively increased mean arterial pressure (MAP) and sympathetic nerve activity. Brain insulin is known to regulate neural activity within the brainstem. We hypothesize that brain insulin injection in T2DM rats attenuates the augmented EPR, and that T2DM is associated with decreased brain insulin. Using male Sprague-Dawley rats, T2DM and control rats were generated via an induction protocol with two low doses of streptozotocin (35 and 25 mg/kg, i.p.) in combination with a 14-23-week high-fat diet or saline injections and a low-fat diet, respectively. After decerebration, MAP and renal sympathetic nerve activity (RSNA) were evaluated during EPR stimulation, evoked by electrically induced muscle contraction via ventral root stimulation, before and after (1 and 2 h post) intracerebroventricular (i.c.v.) insulin microinjections (500 mU, 50 nl). i.c.v. insulin decreased peak MAP (ΔMAP Pre (36 ± 14 mmHg) vs. 1 h (21 ± 14 mmHg) vs. 2 h (11 ± 6 mmHg), P < 0.05) and RSNA (ΔRSNA Pre (107.5 ± 40%), vs. 1 h (75.4 ± 46%) vs. 2 h (51 ± 35%), P < 0.05) responses in T2DM, but not controls. In T2DM rats, cerebrospinal fluid insulin was decreased (0.41 ± 0.19 vs. 0.11 ± 0.05 ng/ml, control (n = 14) vs. T2DM (n = 4), P < 0.01). The results demonstrated that insulin injections into the brain normalized the augmented EPR in brain hypoinsulinaemic T2DM rats, indicating that the EPR can be regulated by brain insulin. KEY POINTS: The reflexive increase in blood pressure and sympathetic nerve activity mediated by the autonomic nervous system during muscle contractions is also known as the exercise pressor reflex. The exercise pressor reflex is dangerously augmented in type 2 diabetes, in both rats and humans. In type 2 diabetic rats both cerebrospinal fluid insulin and phosphoinositide 3-kinase signalling within cardiovascular brainstem neurons decrease in parallel. Brain insulin injections decrease the magnitude of the reflexive pressor and sympathetic responses to hindlimb muscle contraction in type 2 diabetic rats. Partial correction of low insulin within the central nervous system in type 2 diabetes may treat aberrant exercise pressor reflex function.

Blockade of endogenous insulin receptor signaling in the nucleus tractus solitarius potentiates exercise pressor reflex function in healthy male rats.

Insulin not only regulates glucose and/or lipid metabolism but also modulates brain neural activity. The nucleus tractus solitarius (NTS) is a key central integration site for sensory input from working skeletal muscle and arterial baroreceptors during exercise. Stimulation of the skeletal muscle exercise pressor reflex (EPR), the responses of which are buffered by the arterial baroreflex, leads to compensatory increases in arterial pressure to supply blood to working muscle. Evidence suggests that insulin signaling decreases neuronal excitability in the brain, thus antagonizing insulin receptors (IRs) may increase neuronal excitability. However, the impact of brain insulin signaling on the EPR remains fully undetermined. We hypothesized that antagonism of NTS IRs increases EPR function in normal healthy rodents. In decerebrate rats, stimulation of the EPR via electrically induced muscle contractions increased peak mean arterial pressure (MAP) responses 30 min following NTS microinjections of an IR antagonist (GSK1838705, 100 µM; Pre: Δ16 ± 10 mmHg vs. 30 min: Δ23 ± 13 mmHg, n = 11, p = .004), a finding absent in sino-aortic baroreceptor denervated rats. Intrathecal injections of GSK1838705 did not influence peak MAP responses to mechano- or chemoreflex stimulation of the hindlimb muscle. Immunofluorescence triple overlap analysis following repetitive EPR stimulation increased c-Fos overlap with EPR-sensitive nuclei and IR-positive cells relative to sham operation (p < .001). The results suggest that IR blockade in the NTS potentiates the MAP response to EPR stimulation. In addition, insulin signaling in the NTS may buffer EPR stimulated increases in blood pressure via baroreflex-mediated mechanisms during exercise.

Antagonism of TRPV4 channels partially reduces mechanotransduction in rat skeletal muscle afferents.

Mechanical distortion of working skeletal muscle induces sympathoexcitation via thin fibre afferents, a reflex response known as the skeletal muscle mechanoreflex. However, to date, the receptor ion channels responsible for mechanotransduction in skeletal muscle remain largely undetermined. Transient receptor potential vanilloid 4 (TRPV4) is known to sense mechanical stimuli such as shear stress or osmotic pressure in various organs. It is hypothesized that TRPV4 in thin-fibre primary afferents innervating skeletal muscle is involved in mechanotransduction. Fluorescence immunostaining revealed that 20.1 ± 10.1% of TRPV4 positive neurons were small dorsal root ganglion (DRG) neurons that were DiI-labelled, and among them 9.5 ± 6.1% of TRPV4 co-localized with the C-fibre marker peripherin. In vitro whole-cell patch clamp recordings from cultured rat DRG neurons demonstrated that mechanically activated current amplitude was significantly attenuated after the application of the TRPV4 antagonist HC067047 compared to control (P = 0.004). Such reductions were also observed in single-fibre recordings from a muscle-nerve ex vivo preparation where HC067047 significantly decreased afferent discharge to mechanical stimulation (P = 0.007). Likewise, in an in vivo decerebrate rat preparation, the renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) responses to passive stretch of hindlimb muscle were significantly reduced by intra-arterial injection of HC067047 (ΔRSNA: P = 0.019, ΔMAP: P = 0.002). The findings suggest that TRPV4 plays an important role in mechanotransduction contributing to the cardiovascular responses evoked by the skeletal muscle mechanoreflex during exercise. KEY POINTS: Although a mechanical stimulus to skeletal muscle reflexively activates the sympathetic nervous system, the receptors responsible for mechanotransduction in skeletal muscle thin fibre afferents have not been fully identified. Evidence suggests that TRPV4 is a mechanosensitive channel that plays an important role in mechanotransduction within various organs. Immunocytochemical staining demonstrates that TRPV4 is expressed in group IV skeletal muscle afferents. In addition, we show that the TRPV4 antagonist HC067047 decreases the responsiveness of thin fibre afferents to mechanical stimulation at the muscle tissue level as well as at the level of dorsal root ganglion neurons. Moreover, we demonstrate that intra-arterial HC067047 injection attenuates the sympathetic and pressor responses to passive muscle stretch in decerebrate rats. These data suggest that antagonism of TRPV4 attenuates mechanotransduction in skeletal muscle afferents. The present study demonstrates a probable physiological role for TRPV4 in the regulation of mechanical sensation in somatosensory thin fibre muscle afferents.

Exaggerated renal sympathetic nerve and pressor responses during spontaneously occurring motor activity in hypertensive rats.

Stimulation of the mesencephalic locomotor region elicits exaggerated sympathetic nerve and pressor responses in spontaneously hypertensive rats (SHR) as compared with normotensive Wistar-Kyoto rats (WKY). This suggests that central command or its influence on vasomotor centers is augmented in hypertension. The decerebrate animal model possesses an ability to evoke intermittent bouts of spontaneously occurring motor activity (SpMA) and generates cardiovascular responses associated with the SpMA. It remains unknown whether the changes in sympathetic nerve activity and hemodynamics during SpMA are altered by hypertension. To test the hypothesis that the responses in renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) during SpMA are exaggerated with hypertension, this study aimed to compare the responses in decerebrate, paralyzed SHR, WKY, and normotensive Sprague-Dawley (SD) rats. In all strains, an abrupt increase in RSNA occurred in synchronization with tibial motor discharge (an index of motor activity) and was followed by rises in MAP and heart rate. The centrally evoked increase in RSNA and MAP during SpMA was much greater (306 ± 110%) in SHR than WKY (187 ± 146%) and SD (165 ± 44%). Although resting baroreflex-mediated changes in RSNA were not different across strains, mechanically or pharmacologically induced elevations in MAP attenuated or abolished the RSNA increase during SpMA in WKY and SD but had no effect in SHR. It is likely that the exaggerated sympathetic nerve and pressor responses during SpMA in SHR are induced along a central command pathway independent of the arterial baroreflex and/or result from central command-induced inhibition of the baroreflex.

Neural discharge of muscle afferents and pressor response to mechanical stimulation are associated with muscle deformation velocity in rats.

Skeletal muscle reflexes play a crucial role in determining the magnitude of the cardiovascular response to exercise. However, evidence supporting an association between the magnitude of the pressor response and the velocity of muscle deformation has remained to be elucidated. Thus, we investigated the impact of different muscle deformation rates on the neural discharge of muscle afferents and pressor and sympathetic responses in Sprague-Dawley rats. In an ex vivo muscle-nerve preparation, action potentials elicited by sinusoidal mechanical stimuli (137 mN) at different frequencies (0.01, 0.05, 0.1, 0.2, and 0.25 Hz) were recorded in mechanosensitive group III and IV fibers. The afferent response magnitude to sine-wave stimulation significantly varied at different frequencies (ANOVA, P = 0.01). Specifically, as compared with 0.01 Hz (0.83 ± 0.96 spikes/s), the response magnitudes were significantly greater at 0.20 Hz (4.07 ± 5.04 spikes/s, P = 0.031) and 0.25 Hz (4.91 ± 5.30 spikes/s, P = 0.014). In an in vivo decerebrated rat preparation, renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) responses to passive stretch (1 kg) of hindlimb skeletal muscle at different velocities of loading (slow, medium, and fast) were measured. Pressor responses to passive stretch were significantly associated with the velocity of muscle deformation (ANOVA, P < 0.001). The MAP response to fast stretch (Δ 56 ± 12 mmHg) was greater than slow (Δ 33 ± 11 mmHg, P = 0.006) or medium (Δ 30 ± 11 mmHg, P < 0.001) stretch. Likewise, the RSNA response was related to deformation velocity (ANOVA, P = 0.024). These findings suggest that the muscle neural afferent discharge and the cardiovascular response to mechanical stimulation are associated with muscle deformation velocity.

Insulin potentiates the response to capsaicin in dorsal root ganglion neurons in vitro and muscle afferents ex vivo in normal healthy rodents.

Systemic insulin administration evokes sympathoexcitatory actions, but the mechanisms underlying these observations are unknown. We reported that insulin sensitizes the response of thin-fibre primary afferents, as well as the dorsal root ganglion (DRG) that subserves them, to mechanical stimuli. However, little is known about the effects of insulin on primary neuronal responses to chemical stimuli. TRPV1, whose agonist is capsaicin (CAP), is widely expressed on chemically sensitive metaboreceptors and/or nociceptors. The aim of this investigation was to determine the effects of insulin on CAP-activated currents in small DRG neurons and CAP-induced action potentials in thin-fibre muscle afferents of normal healthy rodents. Additionally, we investigated whether insulin potentiates sympathetic nerve activity (SNA) responses to CAP. In whole-cell patch-clamp recordings from cultured mice DRG neurons in vitro, the fold change in CAP-activated current from pre- to post-application of insulin (n = 13) was significantly (P < 0.05) higher than with a vehicle control (n = 14). Similar results were observed in single-fibre recording experiments ex vivo as insulin potentiated CAP-induced action potentials compared to vehicle controls (n = 9 per group, P < 0.05). Furthermore, insulin receptor blockade with GSK1838705 significantly suppressed the insulin-induced augmentation in CAP-activated currents (n = 13) as well as the response magnitude of CAP-induced action potentials (n = 9). Likewise, the renal SNA response to CAP after intramuscular injection of insulin (n = 8) was significantly (P < 0.05) greater compared to vehicle (n = 9). The findings suggest that insulin potentiates TRPV1 responsiveness to CAP at the DRG and muscle tissue levels, possibly contributing to the augmentation in sympathoexcitation during activities such as physical exercise. KEY POINTS: Evidence suggests insulin centrally activates the sympathetic nervous system, and a chemical stimulus to tissues activates the sympathetic nervous system via thin fibre muscle afferents. Insulin is reported to modulate putative chemical-sensitive channels in the dorsal root ganglion neurons of these afferents. In the present study, it is demonstrated that insulin potentiates the responsiveness of thin fibre afferents to capsaicin at muscle tissue levels as well as at the level of dorsal root ganglion neurons. In addition, it is demonstrated that insulin augments the sympathetic nerve activity response to capsaicin in vivo. These data suggest that sympathoexcitation is peripherally mediated via insulin-induced chemical sensitization. The present study proposes a possible physiological role of insulin in the regulation of chemical sensitivity in somatosensory thin fibre muscle afferents.

Insulin potentiates the response to mechanical stimuli in small dorsal root ganglion neurons and thin fibre muscle afferents in vitro.

KEY POINTS: Insulin is known to activate the sympathetic nervous system centrally. A mechanical stimulus to tissues activates the sympathetic nervous system via thin fibre afferents. Evidence suggests that insulin modulates putative mechanosensitive channels in the dorsal root ganglion neurons of these afferents. In the present study, we report the novel finding that insulin augments the mechanical responsiveness of thin fibre afferents not only at dorsal root ganglion, but also at muscle tissue levels. Our data suggest that sympathoexcitation is mediated via the insulin-induced mechanical sensitization peripherally. The present study proposes a novel physiological role of insulin in the regulation of mechanical sensitivity in somatosensory thin fibre afferents. ABSTRACT: Insulin activates the sympathetic nervous system, although the mechanism underlying insulin-induced sympathoexcitation remains to be determined. A mechanical stimulus to tissues such as skin and/or skeletal muscle, no matter whether the stimulation is noxious or not, activates the sympathetic nervous system via thin fibre afferents. Evidence suggests that insulin modulates putative mechanosensitive channels in the dorsal root ganglion (DRG) neurons of these afferents. Accordingly, we investigated whether insulin augments whole-cell current responses to mechanical stimuli in small DRG neurons of normal healthy mice. We performed whole-cell patch clamp recordings using cultured DRG neurons and observed mechanically-activated (MA) currents induced by mechanical stimuli applied to the cell surface. Local application of vehicle solution did not change MA currents or mechanical threshold in cultured DRG neurons. Insulin (500 mU mL-1 ) significantly augmented the amplitude of MA currents (P < 0.05) and decreased the mechanical threshold (P < 0.05). Importantly, pretreatment with the insulin receptor antagonist, GSK1838705, significantly suppressed the insulin-induced potentiation of the mechanical response. We further examined the impact of insulin on thin fibre muscle afferent activity in response to mechanical stimuli in normal healthy rats in vitro. Using a muscle-nerve preparation, we recorded single group IV fibre activity to a ramp-shaped mechanical stimulation. Insulin significantly decreased mechanical threshold (P < 0.05), although it did not significantly increase the response magnitude to the mechanical stimulus. In conclusion, these data suggest that insulin augments the mechanical responsiveness of small DRG neurons and potentially sensitizes group IV afferents to mechanical stimuli at the muscle tissue level, possibly contributing to insulin-induced sympathoexcitation.

Exaggerated pressor and sympathetic responses to stimulation of the mesencephalic locomotor region and exercise pressor reflex in type 2 diabetic rats.

The cardiovascular responses to exercise are potentiated in patients with type 2 diabetes mellitus (T2DM). However, the underlying mechanisms causing this abnormality remain unknown. Central command (CC) and the exercise pressor reflex (EPR) are known to contribute significantly to cardiovascular control during exercise. Thus these neural signals are viable candidates for the generation of the abnormal circulatory regulation in this disease. We hypothesized that augmentations in CC as well as EPR function contribute to the heightened cardiovascular responses during exercise in T2DM. To test this hypothesis, changes in mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA) in response to electrical stimulation of mesencephalic locomotor region (MLR), a putative component of the central command pathway, and activation of the EPR, evoked by electrically induced hindlimb muscle contraction, were examined in decerebrate animals. Sprague-Dawley rats were given either a normal diet (control) or a high-fat diet (14-16 wk) in combination with two low doses (35 mg/kg week 1, 25 mg/kg week 2) of streptozotocin (T2DM). The changes in MAP and RSNA responses to MLR stimulation were significantly greater in T2DM compared with control (2,739 ± 123 vs. 1,298 ± 371 mmHg/s, 6,326 ± 1,621 vs. 1,390 ± 277%/s, respectively, P < 0.05). Similarly, pressor and sympathetic responses to activation of the EPR in diabetic animals were significantly augmented compared with control animals (436 ± 74 vs. 134 ± 44 mmHg/s, 645 ± 135 vs. 139 ± 65%/s, respectively, P < 0.05). These findings provide the first evidence that CC and the EPR may generate the exaggerated rise in sympathetic activity and blood pressure during exercise in T2DM.

Dynamic exercise training prevents exercise pressor reflex overactivity in spontaneously hypertensive rats.

Cardiovascular responses to exercise are exaggerated in hypertension. We previously demonstrated that this heightened cardiovascular response to exercise is mediated by an abnormal skeletal muscle exercise pressor reflex (EPR) with important contributions from its mechanically and chemically sensitive components. Exercise training attenuates exercise pressor reflex function in healthy subjects as well as in heart failure rats. However, whether exercise training has similar physiological benefits in hypertension remains to be elucidated. Thus we tested the hypothesis that the EPR overactivity manifest in hypertension is mitigated by exercise training. Changes in mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA) in response to muscle contraction, passive muscle stretch, and hindlimb intra-arterial capsaicin administration were examined in untrained normotensive Wistar-Kyoto rats (WKYUT; n = 6), exercise-trained WKY (WKYET; n = 7), untrained spontaneously hypertensive rats (SHRUT; n = 8), and exercise-trained SHR (SHRET; n = 7). Baseline MAP after decerebration was significantly decreased by 3 mo of wheel running in SHRET (104 ± 9 mmHg) compared with SHRUT (125 ± 10 mmHg). As previously reported, the pressor and renal sympathetic responses to muscle contraction, stretch, and capsaicin administration were significantly higher in SHRUT than WKYUT. Exercise training significantly attenuated the enhanced contraction-induced elevations in MAP (SHRUT: 53 ± 11 mmHg; SHRET: 19 ± 3 mmHg) and RSNA (SHRUT: 145 ± 32%; SHRET: 57 ± 11%). Training produced similar attenuating effects in SHR during passive stretch and capsaicin administration. These data demonstrate that the abnormally exaggerated EPR function that develops in hypertensive rats is significantly diminished by exercise training.

Exercise training improves functional sympatholysis in spontaneously hypertensive rats through a nitric oxide-dependent mechanism.

Functional sympatholysis is impaired in hypertensive animals and patients. Exercise training (ET) improves functional sympatholysis through a nitric oxide (NO)-dependent mechanism in normotensive rats. However, whether ET has similar physiological benefits in hypertension remains to be elucidated. Thus we tested the hypothesis that the impairment in functional sympatholysis in hypertension is reversed by ET through a NO-dependent mechanism. In untrained normotensive Wistar-Kyoto rats (WKYUT; n = 13), untrained spontaneously hypertensive rats (SHRUT; n = 13), and exercise-trained SHR (SHRET; n = 6), changes in femoral vascular conductance (FVC) were examined during lumbar sympathetic nerve stimulation (1, 2.5, and 5 Hz) at rest and during muscle contraction. The magnitude of functional sympatholysis (Δ%FVC = Δ%FVC muscle contraction - Δ%FVC rest) in SHRUT was significantly lower than WKYUT (1 Hz: -2 ± 4 vs. 13 ± 3%; 2.5 Hz: 9 ± 3 vs. 21 ± 3%; and 5 Hz: 12 ± 3 vs. 26 ± 3%, respectively; P < 0.05). Three months of voluntary wheel running significantly increased maximal oxygen uptake in SHRET compared with nontrained SHRUT (78 ± 6 vs. 62 ± 4 ml·kg(-1)·min(-1), respectively; P < 0.05) and restored the magnitude of functional sympatholysis in SHRET (1 Hz: 9 ± 2%; 2.5 Hz: 20 ± 4%; and 5 Hz: 34 ± 5%). Blockade of NO synthase (NOS) by N(G)-nitro-l-arginine methyl ester attenuated functional sympatholysis in WKYUT but not SHRUT. Furthermore, NOS inhibition significantly diminished the improvements in functional sympatholysis in SHRET. These data demonstrate that impairments in functional sympatholysis are normalized via a NO mechanism by voluntary wheel running in hypertensive rats.

A role for nitric oxide within the nucleus tractus solitarii in the development of muscle mechanoreflex dysfunction in hypertension.

Evidence suggests that the muscle mechanoreflex, a circulatory reflex that raises blood pressure and heart rate (HR) upon activation of mechanically sensitive afferent fibres in skeletal muscle, is overactive in hypertension. However, the mechanisms underlying this abnormal reflex function have yet to be identified. Sensory input from the mechanoreflex is processed within the nucleus tractus solitarii (NTS) in the medulla oblongata. Within the NTS, the enzymatic activity of nitric oxide synthase produces nitric oxide (NO). This centrally derived NO has been shown to modulate muscle reflex activity and serves as a viable candidate for mediating the mechanoreflex dysfunction that develops in hypertension. We hypothesized that mechanoreflex dysfunction in hypertension is mediated by abnormal alterations in NO production in the NTS. Mechanically sensitive afferent fibres were stimulated by passively stretching hindlimb muscle before and after blocking the endogenous production of NO within the NTS via microdialysis of the NO synthase inhibitor L-NAME (1 and 5 mM) in normotensive Wistar-Kyoto rats and spontaneously hypertensive rats (SHRs). Changes in HR and mean arterial pressure in response to stretch were significantly larger in SHRs compared with Wistar-Kyoto rats prior to L-NAME dialysis. Attenuating NO production via L-NAME in normotensive rats recapitulated the exaggerated cardiovascular response to stretch observed in SHRs. Dialysing L-NAME in SHRs further accentuated the increases in HR and mean arterial pressure elicited by stretch. These findings support the contention that reductions in NO production within the NTS contribute to the generation of abnormal cardiovascular control by the skeletal muscle mechanoreflex in hypertension.

TRPV1 (Transient Receptor Potential Vanilloid 1) Sensitization of Skeletal Muscle Afferents in Type 2 Diabetic Rats With Hyperglycemia.

[Figure: see text].

Skeletal Muscle Reflex-Induced Sympathetic Dysregulation and Sensitization of Muscle Afferents in Type 1 Diabetic Rats.

The blood pressure response to exercise is exaggerated in the type 1 diabetes mellitus (T1DM). An overactive exercise pressor reflex (EPR) contributes to the potentiated pressor response. However, the mechanism(s) underlying this abnormal EPR activity remains unclear. This study tested the hypothesis that the heightened blood pressure response to exercise in T1DM is mediated by EPR-induced sympathetic overactivity. Additionally, the study examined whether the single muscle afferents are sensitized by PKC (protein kinase C) activation in this disease. Sprague-Dawley rats were intraperitoneally administered either 50 mg/kg streptozotocin (T1DM) or saline (control). At 1 to 3 weeks after administration, renal sympathetic nerve activity and mean arterial pressure responses to activation of the EPR, mechanoreflex, and metaboreflex were measured in decerebrate animals. Action potential responses to mechanical and chemical stimulation were determined in group IV afferents with pPKCα (phosphorylated-PKCα) levels assessed in dorsal root ganglia. Compared with control, EPR (58±18 versus 96±33%; P<0.05), mechanoreflex (21±13 versus 51±20%; P<0.05), and metaboreflex (40±20 versus 88±39%; P<0.01) activation in T1DM rats evoked significant increases in renal sympathetic nerve activity as well as mean arterial pressure. The response of group IV afferents to mechanical (18±24 versus 61±45 spikes; P<0.01) and chemical (0.3±0.4 versus 1.6±0.8 Hz; P<0.01) stimuli were significantly greater in T1DM than control. T1DM rats showed markedly increased pPKCα levels in dorsal root ganglia compared with control. These data suggest that in T1DM, abnormally muscle reflex-evoked increases in sympathetic activity mediate exaggerations in blood pressure. Further, sensitization of muscle afferents, potentially via PKC activation, may contribute to this abnormal circulatory responsiveness.

Insulin resistance is associated with an exaggerated blood pressure response to ischemic rhythmic handgrip exercise in nondiabetic older adults.

Patients with type 2 diabetes display an exaggerated pressor response to exercise. However, evidence supporting the association between the magnitude of the pressor response to exercise and insulin resistance-related factors including hemoglobin A1c (HbA1c) or homeostatic model assessment of insulin resistance (HOMA-IR) in nondiabetic subjects has remained sparse and inconclusive. Thus we investigated the relationship between cardiovascular responses to exercise and insulin resistance-related factors in nondiabetic healthy men (n = 23) and women (n = 22) above 60 yr old. We measured heart rate (HR) and blood pressure (BP) responses during: isometric handgrip (IHG) exercise of 30% maximal voluntary contraction, a period of skeletal muscle ischemia (SMI) induced by tourniqueting the arm after IHG, and rhythmic dynamic handgrip (DHG) exercise during SMI. Greater diastolic BP (DBP) responses to DHG with SMI was associated with male sex (r = 0.44, P = 0.02) and higher HbA1c (r = 0.33, P = 0.03), heart-ankle pulse wave velocity (haPWV) (r = 0.45, P < 0.01), and resting systolic BP (SBP) (r = 0.36, P = 0.02). HbA1c persisted as a significant determinant explaining the variance in the DBP response to DHG with SMI in multivariate models despite adjustment for sex, haPWV, and resting SBP. It was also determined that the DBP response to DHG with SMI in a group in which HOMA-IR was abnormal (Δ33 ± 3 mmHg) was significantly higher than that of groups in which HOMA-IR was at intermediate (Δ20 ± 4 mmHg) and normal (Δ23 ± 2 mmHg) levels. These data suggest that even in nondiabetic older adults, insulin resistance is related to an exaggerated pressor response to exercise especially when performed under ischemic conditions.NEW & NOTEWORTHY The diastolic blood pressure response to rhythmic dynamic handgrip exercise under ischemic conditions was demonstrated to be correlated with insulin resistance-related factors in nondiabetic older adults. This finding provides important insight to the prescription of exercise in this particular patient population as the blood pressure response to exercise, especially under ischemic conditions, could be exaggerated to nonsafe levels.

The Pressor Response to Concurrent Stimulation of the Mesencephalic Locomotor Region and Peripheral Sensory Afferents Is Attenuated in Normotensive but Not Hypertensive Rats.

Central command (CC) and the exercise pressor reflex (EPR) regulate blood pressure during exercise. We previously demonstrated that experimental stimulation of the CC and EPR pathways independently contribute to the exaggerated pressor response to exercise in hypertension. It is known that CC and EPR modify one another functionally. Whether their interactive relationship is altered in hypertension, contributing to the generation of this potentiated blood pressure response, remains unknown. To address this issue, the pressor response to activation of the CC pathway with and without concurrent stimulation of the EPR pathway, and vice versa, was examined in normotensive Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) rats. In decerebrated, paralyzed animals, activation of the CC pathway was evoked by electrical stimulation of the mesencephalic locomotor region (MLR; 20-50 µA in 10-µA steps). Electrical stimulation of the sciatic nerve (SN, 3, 5, and 10 × motor threshold; MT) was used to activate hindlimb afferents known to carry EPR sensory information. In both WKY and SHR, the algebraic sum of the pressor responses to individual stimulation of the MLR and SN were greater than when both inputs were stimulated simultaneously. Although the blood pressure response to a constant level of SN stimulation was not significantly affected by concurrent MLR stimulation at variable intensities, the pressor response to a constant level of MLR simulation was significantly attenuated by concurrent SN stimulation in WKY but not in SHR. These findings suggest the interactive relationship between CC and the EPR is inhibitory in nature in both WKY and SHR. However, the neural occlusion between these central and peripheral pressor mechanisms is attenuated in hypertension.

Impact of neonatal hypothyroidism on reproduction in the male hamster, Mesocricetus auratus.

Thyroid hormones play an important role in brain development. In the present study, we examined the influence of transient postnatal hypothyroidism on reproductive neuroendocrine and behavioral outcomes in the male Syrian (golden) hamster. Hamster pups were rendered hypothyroid following exposure to the goitrogen, 6-n-propyl-2-thiouracil (PTU), between postnatal (PN) day 0 (birth) and PN25 (weaning). By 15 days after cessation of PTU, exposure (PN40) serum thyroxine levels had returned to control levels. The testes of treated males were approximately 30% heavier than controls and daily sperm production was increased by 73%. Immunocytochemistry for GnRH revealed that the total number of GnRH neurons did not vary between groups; however, a shift in the distribution of GnRH neurons was observed in treated males such that more GnRH immunoreactive neurons were found in the caudal portion of their normal distribution. The shift in GnRH distribution was associated with a significant reduction (40-50%) in pituitary gonadotropin secretion. Behaviorally, treated males took significantly longer to investigate the anogenital region and then mount a receptive female. A corresponding reduction in the total number of anogenital investigations and mounts was observed. This difference between treated males and controls was reduced, but not eliminated, over successive trials and by the third trial the number of intromission was similar between treated and control males. We conclude that the full complement of adult reproductive functions observed in the male golden hamster requires thyroid hormones during the early postnatal period. The severity of the effects induced by early hypothyroidism in this species varies from transient to permanent, depending on the endpoint.

Salicylate-based analgesic balm attenuates pressor responses from skeletal muscle.

PURPOSE: Analgesic balms (AB) are widely used in sports medicine. We previously have examined effects of various counterirritant-based AB on pressor responses evoked by muscular contraction (MC), mediated by group III and IV muscle afferents known to produce exercise and nociceptive responses. Our purpose was to examine trolamine salicylate-based analgesic balm (TS) effects. METHODS: Ten healthy, adult male and female cats were used. Decerebration under halothane allowed elimination of anesthesia. Electrical stimulation of L7 and S1 ventral roots evoked static MC (30 s). After control runs, commercial TS (10% concentration) was applied to the skin over the contracting muscles of one hind limb (N = 5). MC was evoked every 10 min, alternating between sides. Ipsilateral (T = 0, T + 20, T + 40, T + 60 min) and contralateral (T - 10, T + 10, T + 30, T + 50 min) responses were analyzed. Five additional cats received AB minus TS. RESULTS: There were significant attenuations in both peak mean arterial pressure (MAP), in the last 12 s and the last 6 s of the 30 s of MC for both contra- and ipsilateral limbs occurring at T + 50 and T + 60 min after TS application, respectively. No significant changes in heart rate (HR) responses were seen for either the ipsi- or contralateral stimulation. There were no changes in MAP or HR in control cats. CONCLUSIONS: These results indicate that TS affects the end of the 30 s of MC, which is thought to be mainly chemically mediated through group IV afferents. TS represents the salicylate class of AB and has no counterirritant properties. TS works as an inhibitor of cyclooxygenase (prostaglandin formation) and is, at least in part, blood borne.

Reversibility of exercise-induced dendritic attenuation in brain cardiorespiratory and locomotor areas following exercise detraining.

It has been shown previously that dendritic branching in cardiorespiratory and locomotor brain areas can be attenuated with exercise training (ET). It was not known whether this process was reversible. Twenty-three (n = 23) male Sprague-Dawley rats were individually caged and divided into two groups: untrained (UN; n = 11) and detrained (DTR; n = 12). DTR were provided with a running wheel at 21 days of age and exercised spontaneously. After 120 days (70 days of ET followed by 50 days of detraining), ET indexes were obtained, including maximal oxygen uptake, percent body fat, resting heart rate, and heart weight-to-body weight ratios. The brain was processed according to a modified Golgi-Cox procedure. Impregnated neurons from the periaqueductal gray (PAG), posterior hypothalamic area (PH), nucleus of the tractus solitarius (NTS), and cuneiform nucleus (CfN) were examined in coronal sections. Neurons were traced using a camera lucida technique and analyzed using the Sholl concentric ring analysis of dendritic branching. t-Tests compared the mean number of intersections per neuron by grouping inner rings, outer rings, and total number of intersections per animal. There were no significant differences between UN and DTR in PH, PAG, CfN, and NTS in the inner rings, outer rings, and total number of intersections per animal. A separate group of animals was used to show that a training effect in the CfN and NTS was present at 56 days of ET. Our results show that dendritic attenuation resulting from 70 days of ET in PH, PAG, CfN, and NTS is completely reversed with 50 days of detraining.

Loss of cyclin-dependent kinase inhibitors produces adipocyte hyperplasia and obesity.

Adipocyte hyperplasia is characteristic of some forms of human obesity, but the role of adipocyte number in obesity and how normal adipocyte number is established are unclear. Preadipocytes proliferate and then differentiate to become mitotically quiescent adipocytes. This involves exit from the cell cycle, a process regulated by cell cycle inhibitors such as the cyclin-dependent kinase inhibitors (CDKIs) p27 and p21. 3T3-L1 preadipocytes show marked changes in p27 and p21 during differentiation, suggesting CDKIs may regulate establishment of adipocyte number in vivo. To study the role of these CDKIs in adipogenesis, we analyzed adult p27 knockout (p27KO), p21 knockout (p21KO), p27/p21 double knockout (DBKO), and wild-type (WT) mice. Adult DBKO mice weighed 100% more and had fourfold increases in body fat percentage compared with WT. Fat pad weights were increased 80, 90, and 500% in p27KO, p21KO, and DBKO mice, respectively, compared with WT. Adipocyte numbers of p27KO, p21KO, and DBKO mice were 1.9-, 1.7-, and 6.1-fold, respectively, that of WT; adipocyte size was not increased. DBKO mice showed glucose intolerance, insulin insensitivity, hepatic steatosis and dyslipidemia; gradations of these effects occurred in p27KO and p21KO mice. In conclusion, p27KO and p21KO mice are obese because of adipocyte hyperplasia, and DBKO mice have further increases in obesity and adipocyte hyperplasia, indicating that their functions in establishing adipocyte number are not redundant. p27 and p21 are major regulators of adipocyte number in vivo, and knockouts lacking one or both of these proteins provide models for producing adipocyte hyperplasia and understanding its metabolic consequences.

Neuroplastic adaptations to exercise: neuronal remodeling in cardiorespiratory and locomotor areas.

Neuronal activity has been shown to be attenuated in cardiorespiratory and locomotor centers of the brain in response to a single bout of exercise in trained (TR) vs. untrained (UN) animals, but the mechanisms remain obscure. Based on this finding, dendritic branching patterns of seven brain areas associated with cardiorespiratory and locomotor activity were examined in TR and UN animals. Twenty-eight male Sprague-Dawley rats were kept in individual cages and divided into TR and UN. TR were provided with a running wheel and exercised spontaneously. After 85 or 120 days, exercise training indexes were obtained, including maximal oxygen consumption, percent body fat, resting heart rate, and heart weight-to-body weight ratios. The brain was removed and processed according to a modified Golgi-Cox procedure. Impregnated neurons from seven brain areas were examined in coronal sections: the periaqueductal gray, posterior hypothalamic area, nucleus of the tractus solitarius, rostral ventrolateral medulla, cuneiform nucleus, nucleus cuneatus, and cerebral cortex. Neurons were traced using a camera lucida technique and analyzed using the Sholl analysis of dendritic branching. t-tests were conducted to compare the mean number of intersections per neuron by grouping inner rings and outer rings and also comparing the total number of intersections per animal. There were significant differences between groups in the posterior hypothalamic area, periaqueductal gray, cuneiform nucleus, and nucleus of the tractus solitarius in the inner rings, outer rings, and the total number of intersections per animal. Our results show that dendritic fields of neurons in important cardiorespiratory and locomotor centers of the brain are attenuated in TR animals.