Regulation Of Sympathetic Nerve Discharge Research Articles

Does Acute Heat Stress Differentially‐Modulate RVLM Gene Expression of Ionotropic Neurotransmitter Receptors in Young and Aged F344 Rats?

The rostral ventrolateral medulla (RVLM) is involved in the regulation of sympathetic nerve discharge (SND) and arterial blood pressure. Acute hyperthermia produces marked activation of visceral SND in young rats, and the functional integrity of this brainstem region is crucial for sustaining visceral sympathoexcitation to increased internal body temperature (Tc). Visceral SND responses to increased Tc are attenuated in aged compared with young F344 rats, and central mechanisms contributing to age‐associated alterations in SND responses to hyperthermia are not well‐established. The balance between excitatory and inhibitory inputs, mediated primarily by glutamate and GABA neural systems, is important for regulating RVLM neuronal activity. In the present study we hypothesized that, in response to acute hyperthermia, the RVLM gene expression of various excitatory and inhibitory ionotropic neurotransmitter receptors would demonstrate age‐related differences. Tc was increased from 38°C to 41.5°C in young (3–4 months) and aged (23–24 months) anesthetized F344 rats. Total RNA was extracted from RVLM‐containing micropunches and gene expression analysis for ionotropic neurotransmitter receptor subunits was performed using TaqMan real time RT‐PCR. Gene expression levels of glutamate, GABA and glycine neurotransmitter binding receptor subunits were compared between age groups. Gene expression analysis revealed an upregulation in some, but not all, major excitatory NMDA and AMPA neurotransmitter receptor subunits in young compared with aged rats. In addition, major subunits of the inhibitory GABA and glycine receptors demonstrated substantial upregulation in aged compared with young rats. These preliminary data suggest that, in response to acute hyperthermia, aging modulates the functional balance of RVLM excitatory and inhibitory neurotransmitter receptor subunits.Support or Funding InformationNIH AG‐041948.

Analysis of RVLM Gene Expression in Young, Middle‐Aged and Aged F344 Rats

Abstract: The rostral ventral lateral medulla (RVLM) plays a critical role in regulation of basal sympathetic nerve discharge (SND). Advanced age is associated with altered regulation of SND, however, the effect of age on central medullary mechanisms regulating SND remains unknown. A series of experiments were initiated to identify cellular and molecular targets that may mediate age‐related changes in medullary regulation of SND. We tested the hypothesis that aging shifts the balance of RVLM excitatory and inhibitory influences towards excitation. Gene expression analysis was performed using Agilent Whole Rat Genome Microarray 4×44K followed by validation of microarray with TaqMan real‐time RT‐PCR and immunoblotting for specific RVLM proteins. Total RNA and protein were extracted from RVLM micropunches from young, middle‐aged and aged F344 rats. Microarray analysis revealed up‐regulation in aged compared to young and middle‐aged rats of several genes related to immune function and neurotransmitter receptors. Specifically, the chemokines CCL3, CCL4 and CCL5 were up‐regulated 3 to 4 fold in aged rats compared to young and middle‐aged rats. The results of previous studies suggest that increased chemokines in the brain can modulate NMDA receptor responses. The current preliminary results suggest that increased chemokine gene expression in the RVLM of aged rats may contribute to age‐related changes in SND regulation. Supported by NIH A6‐041948.

Dexmedetomidine and regulation of splenic sympathetic nerve discharge in aged F344 rats.

Sedatives influence the immune system and centrally-acting alpha2-adrenergic receptor agonists, including Dexmedetomidine (Dex), modulate sympathetic nerve discharge (SND). Because sedatives are used under medical conditions that include elderly patients, and because advancing age attenuates SND responsivity to various interventions, we tested the hypothesis that splenic sympathoinhibitory responses to Dex would be attenuated in aged compared with young Fischer 344 rats. Dex-mediated reductions in splenic SND were similar in aged and young baroreceptor-intact and -denervated rats, indicating that SND changes to Dex administration occur in an age-independent manner. These findings provide new information regarding interactions between alpha2-adrenergic agonists, advanced age, and SND regulation.

Disinhibition of RVLM neural circuits and regulation of sympathetic nerve discharge at peak hyperthermia

Hyperthermia is a potent activator of visceral sympathetic nerve discharge (SND), and the functional integrity of the rostral ventral lateral medulla (RVLM) is critically important for sustaining sympathoexcitation at peak hyperthermia. However, RVLM mechanisms mediating SND activation to acute heat stress are not well understood. Because RVLM GABA is tonically inhibitory to sympathetic nerve outflow, it is plausible to hypothesize that disinhibition of RVLM sympathetic neural circuits, via withdrawal of GABAergic tone, may affect SND regulation at peak hyperthermia. The effect of RVLM bicuculline (BIC; GABAA receptor antagonist, 100-200 pmol) microinjections on the level of renal SND in anesthetized rats was determined after internal body temperature (Tc) had been increased to 41.5°C. Temperature-control experiments involved RVLM BIC (100-200 pmol) microinjections, with Tc maintained at 38°C. As expected, acute heating significantly increased renal SND from control levels. Bilateral RVLM BIC microinjections at 41.5°C produced immediate and significant increases in renal SND above heating-induced levels of activation. Bilateral RVLM BIC microinjections at 38°C increased renal SND to similar levels as produced by RVLM BIC microinjections after Tc had been increased to 41.5°C (heating + RVLM BIC). These results demonstrate that a considerable level of RVLM GABAergic inhibition is sustained at peak hyperthermia, an interesting physiological response profile based on the significance of SND activation to cardiovascular regulation during heat stress.

Translational physiology and SND recordings in humans and rats: A glimpse of the recent past with an eye on the future

The sympathetic nervous system (SNS) plays an important role in cardiovascular function, and based on the critical mechanistic relationship between altered sympathetic neural mechanisms and cardiovascular disease, it is important that the autonomic research community identifies deficiencies in the translational exchange of information and strives for a more thorough understanding of the translational significance of findings from studies involving sympathetic nerve discharge (SND) regulation in human and animal subjects. The present review assesses the state of the literature regarding studies that have used direct recordings of SND during the past three decades in humans and rats, focusing on; 1) identifying the number of studies reporting SND recordings in humans and rats, 2) briefly describing the translational exchange of SND regulation information from these studies, 3) contrasting the number of studies completed in anesthetized and conscious rats, and 4) assessing the prevalence of long-term SND recording studies in conscious rats. The majority of SND recordings in rats have been completed using anesthetized preparations, although a substantial number of studies have been completed in conscious rats. However, few studies have completed long-term (>5 days) SND recordings in freely-behaving rats, and even fewer studies have used experimental preparations that combine long-term nerve recordings with the capacity for completing central neural microinjections, or have been completed in animal models of cardiovascular disease. The wide-spread implementation of long-term SND recordings in rodent models of cardiovascular disease would be expected to enhance the translational exchange of clinically-relevant information between animals and humans.

Effects of combined aging and heart failure on visceral sympathetic nerve and cardiovascular responses to progressive hyperthermia in F344 rats

Sympathetic nerve discharge (SND) responses to hyperthermia are attenuated in aged rats without heart failure (HF) and in young HF (Y(HF)) rats, demonstrating that individually aging and HF alter SND regulation. However, the combined effects of aging and HF on SND regulation to heat stress are unknown, despite the high prevalence of HF in aged individuals. We hypothesized that SND responses to heating would be additive when aging and HF are combined, demonstrated by marked reductions in SND and mean arterial pressure (MAP) responses to heating in aged HF (A(HF)) compared with aged sham HF (A(SHAM)) rats, and in A(HF) compared with Y(HF) rats. Renal and splenic SND responses to hyperthermia (colonic temperature increased to 41.5°C) were determined in anesthetized Y(HF), young sham (Y(SHAM)), A(HF), and A(SHAM) Fischer rats. HF was induced by myocardial infarction and documented using echocardiographic, invasive, and postmortem measures. The severity of HF was similar in Y(HF) and A(HF) rats. SND responses to heating were attenuated in Y(HF) compared with Y(SHAM) rats, demonstrating an effect of HF on SND regulation in young rats. In contrast, A(HF) and A(SHAM) rats demonstrated similar SND responses to heating, suggesting a prominent influence of age on SND regulation in A(HF) rats. Splenic SND and MAP responses to heating were similar in Y(HF), A(HF), and A(SHAM) rats, indicating that the imposition of HF in young rats changes the regulatory status of these variables to one consistent with aged rats. These data suggest that the effect of HF on SND regulation to hyperthermia is age dependent.

Animal aging and regulation of sympathetic nerve discharge

Studies completed in human subjects have made seminal contributions to understanding the effects of age on sympathetic nervous system (SNS) regulation. Numerous experimental constraints limit the design of studies involving human subjects; therefore, completion of studies in animal models of aging would be expected to provide additional insight regarding mechanisms mediating age-related changes in sympathetic nerve discharge (SND) regulation. The present review assesses the current state of the literature regarding contributions from animal studies on the effects of advancing age on SND regulation, focusing primarily on studies that have used direct recordings of sympathetic nerve outflow. Few studies using direct SND recordings have been completed in animal models of aging, regardless of the fundamental component of SND regulation reviewed (basal levels, acute responsiveness, relationships between the discharges in sympathetic nerves, central neural regulation). SNS responsiveness to various acute stressors is altered in aged compared with young animals; however, mechanisms remain virtually unexplored. There is a marked dearth of studies that have used central neural microinjection techniques in conjunction with SND recordings in aged animals, making it difficult to develop an evidence-based framework regarding potential age-associated effects on central regulation of SND. Determination of age-related changes in mechanisms regulating SND is important for understanding relationships between chronic disease development and changes in SNS function; however, this can only be achieved by substantially extending the current knowledge base regarding the effects of age on SND regulation in animal studies.

Are Aging and Heart Failure Similar Syndromes of Sympathetic Dysregulation?

Both heart failure (HF) and aging alter the basal regulation of sympathetic nerve discharge (SND) and the majority of HF patients are aged, however, the effect of combined aging and HF on SND regulation to acute stress is not well established. We tested the hypothesis that aging and HF are not similar syndromes of sympathetic nervous system dysregulation, as evidenced by differences in SND (renal and splenic) responses to heat stress in young HF compared with aged HF rats and in aged HF compared with aged non‐HF rats. Anesthetized young (3–6 months) and aged (24–25 months) F344 rats received either a surgically‐induced myocardial infarction or a sham operation. Heating experiments (internal body temperature, Tc, increased to 41.5°C) were completed in chloralose‐urethane anesthetized rats 4–6 weeks after the myocardial (or sham) infarction procedure. Left ventricular (LV) function was determined by measuring left ventricular end diastolic pressure (LVEDP) and comparing dimensions of the left atrium (LA) and the aorta (Ao) (LA/Ao ratio) using echocardiographic techniques. Preliminary results indicate that LVEDP and LA/Ao ratio and SND responses to acute thermal stress may differ in young HF compared with aged HF rats and in aged HF compared with aged non‐HF rats. These findings suggest that both age and HF should be considered when determining mechanisms regulating SND responses to acute environmental stress. Funding provided by NIH HL091342.

Central nervous system administration of interleukin-6 produces splenic sympathoexcitation

Interleukin-6 (IL-6) is a multifunctional cytokine that has been shown to play a pivotal role in centrally-mediated physiological responses including activation of the hypothalamic–pituitary–adrenal axis. Cerebral spinal fluid (CSF) concentrations of IL-6 are elevated in multiple pathophysiological conditions including Alzheimer's disease, autoimmune disease, and meningitis. Despite this, the effect of IL-6 on central regulation of sympathetic nerve discharge (SND) remains unknown which limits understanding of sympathetic-immune interactions in health and disease. In the present study we determined the effect of intracerebroventricular (icv, lateral ventricle) administration of IL-6 on splenic SND in urethane–chloralose-anesthetized rats. A second goal was to determine if icv injected IL-6 enters the brain parenchyma and acts as a volume transmission signal to access areas of the brain involved in regulation of sympathetic nerve outflow. Icv administration of IL-6 (10 ng, 100 ng, and 400 ng) significantly and progressively increased splenic SND from control levels in baroreceptor-denervated Sprague–Dawley rats. Administration of 100-ng and 400-ng IL-6 resulted in significantly higher SND responses when compared to those elicited with a 10-ng dose. Sixty minutes following icv administration, fluorescently labeled IL-6 was not distributed throughout the parenchyma of the brain but was localized to the periventricular areas of the ventricular system. Brain sections counter-stained for the IL-6 receptor (IL-6R) revealed that IL-6 and the IL-6R were co-localized in periventricular areas adjoining the third ventricle. These results demonstrate that icv IL-6 administration increases splenic SND, an effect likely achieved via signaling mechanisms originating in the periventricular cells.

Paraventricular Nucleus Interleukin‐6 Receptor Expression in Heart Failure

Heart failure (HF) increases sympathetic nerve discharge (SND) and plasma interleukin 6 (IL-6) levels, which directly correlate with disease severity. Previous research has shown increased mRNA levels of the interleukin-6 receptor (IL-6R) within the paraventricular nucleus (PVN), a forebrain nucleus that integrates autonomic and endocrine signals. The central actions of the immunoregulatory cytokine IL-6 are highly dependent upon the cytokine binding the translated form of the IL-6R. The current study tested the hypothesis that PVN expression of the IL-6R complex is enhanced in HF compared to sham-operated controls. Rats underwent a left-coronary artery ligation (n=15) or sham operation (n=8). Eight weeks after surgery hemodynamic measurements were taken, blood samples collected, brain tissue harvested, and protein analysis performed. Brains were cut in the coronal plane and bilateral punches of PVN and non-PVN sites were taken. Left ventricular end-diastolic pressure (LVEDP) readings were used to divide HF rats into severe (LVEDP ≥ 20 mm Hg) and moderate (LVEDP <20 mm Hg) HF groups. Consistent with previous findings, plasma levels of IL-6 showed a direct correlation with severity of HF. PVN protein expression of IL-6R increased with the severity of HF whereas expression of the beta subunit protein, gp130, was independent of HF severity. These preliminary results suggest that the central expression of the IL-6R is upregulated in HF rats. This upregulation provides a possible path by which IL-6 may alter SND regulation during HF. Supported by NHLBI HL 69755-04, HL 065346-04, and AHA 0520105Z

GABAB-receptor-mediated suppression of sympathetic outflow from the spinal cord of neonatal rats

Using a splanchnic nerve-spinal cord preparation in vitro that could spontaneously generate sympathetic nerve discharge (SND), we investigated the roles of intraspinal GABA(B) receptors in the regulation of SND. Despite an age-dependent difference in sensitivity, bath applications of baclofen (Bac; GABA(B)-receptor agonist) consistently reduced SND in a concentration-dependent manner. The drug specificity of Bac in activation of GABA(B) receptors was verified by application of its antagonist saclofen (Sac) or CGP-46381 (CGP). Sac or CGP alone did not change SND. However, in the presence of Sac or CGP, the effects of Bac on SND inhibition were reversibly attenuated. The splanchnic sympathetic preganglionic neuron (SPN) was recorded by blind whole cell, patch-clamp techniques. We examined Bac effects on electrical membrane properties of SPNs. Applications of Bac reduced excitatory synaptic events, induced membrane hyperpolarizations, and inhibited SPN firing. In the presence of 12 mM Mg2+ or 0.5 microM TTX to block Ca2+- or action potential-dependent synaptic transmissions, applications of Bac induced an outward baseline current that reversed at -29 +/- 6 mV. Because the K+ equilibrium potential in our experimental conditions was -100 mV, the Bac-induced currents could not simply be attributed to an alteration of K+ conductance. On the other hand, applications of Bac to Cs+-loaded SPNs reduced Cd2+-sensitive and high-voltage-activated inward currents, indicating an inhibition of voltage-gated Ca2+ currents. Our results suggest that the activation of intraspinal GABA(B) receptors suppresses SND via a mixture of ion events that may link to a change in Ca2+ conductance.

Central interleukin-1beta antibody increases renal and splenic sympathetic nerve discharge.

We tested the hypothesis that intracerebroventricular (lateral ventricle) administration of interleukin-1beta (IL-1beta) antibody increases the level of sympathetic nerve discharge (SND) in alpha-chloralose-anesthetized rats. Mean arterial pressure (MAP), heart rate (HR), and SND (splenic and renal) were recorded before (Preinfusion), during (25 min), and for 45 min after infusion of IL-1beta antibody (15 microg, 50 microl icv) in baroreceptor-intact (intact) and sinoaortic-denervated (SAD) rats. The following observations were made. First, intracerebroventricular infusion of IL-1beta antibody (but not saline and IgG) significantly increased MAP and the pressor response was higher in SAD compared with intact rats. Second, renal and splenic SND were significantly increased during and after intracerebroventricular IL-1beta antibody infusion and sympathoexcitatory responses were higher in SAD compared with intact rats. Third, intracerebroventricular administration of a single dose of IL-1beta antibody (15 microg, 5 microl for 2 min) significantly increased splenic and renal SND in intact rats. These results suggest that under the conditions of the present experiments central neural IL-1beta plays a role in the tonic regulation of SND and arterial blood pressure.

Role of paraventricular nucleus in regulation of sympathetic nerve frequency components.

Autospectral and coherence analyses were used to determine the role of and interactions between paraventricular nucleus (PVN) nitric oxide, gamma-aminobutyric acid (GABA), and the N-methyl-D-aspartic acid (NMDA)-glutamate receptor in regulation of sympathetic nerve discharge (SND) frequency components in anesthetized rats. Four observations were made. First, PVN microinjection of bicuculline (BIC) (GABA(A) receptor antagonist), but not single PVN injections of NMDA (excitatory amino acid) or N(G)-monomethyl-L-arginine (L-NMMA; a nitric oxide synthase inhibitor), altered SND frequency components. Second, combined PVN microinjections of L-NMMA and NMDA changed the SND bursting pattern; however, the observed pattern change was different from that produced by PVN BIC and not observed after sinoaortic denervation. Third, PVN microinjection of kynurenic acid prevented and reversed BIC-induced changes in the SND bursting pattern. Finally, vascular resistance (renal and splenic) was significantly increased after PVN BIC microinjection despite the lack of change in the level of renal and splenic SND. These data demonstrate that the PVN contains the neural substrate for altering SND frequency components and suggest complex interactions between specific PVN neurotransmitters and between PVN neurotransmitters and the arterial baroreceptor reflex in SND regulation.

Altered frequency responses of sympathetic nerve discharge bursts after IL-1beta and mild hypothermia.

Although interleukin-1beta (IL-1beta) administration produces nonuniform changes in the level of sympathetic nerve discharge (SND), the effect of IL-1beta on the frequency-domain relationships between discharges in different sympathetic nerves is not known. Autospectral and coherence analyses were used to determine the effect of IL-1beta and mild hypothermia (60 min after IL-1beta, colonic temperature from 38 degrees C to 36 degrees C) on the relationships between renal-interscapular brown adipose tissue (IBAT) and splenic-lumbar sympathetic nerve discharges in chloralose-anesthetized rats. The following observations were made. 1) IL-1beta did not alter renal-IBAT coherence values in the 0- to 2-Hz frequency band or at the cardiac frequency (CF). 2) Peak coherence values relating splenic-lumbar discharges at the CF were significantly increased after IL-1beta and during hypothermia. 3) Hypothermia after IL-1beta significantly reduced the coupling (0-2 Hz and CF) between renal-IBAT but not splenic-lumbar SND bursts. 4) Combining IL-1beta and mild hypothermia had a greater effect on renal-IBAT SND coherence values than did mild hypothermia alone. These data demonstrate functional plasticity in sympathetic neural circuits and suggest complex relationships between immune products and SND regulation.

Interleukin-1 beta alters brown adipose tissue but not renal sympathetic nerve responses to hypothermia.

Proinflammatory cytokines and acute physical stress influence sympathetic nerve discharge (SND). Because interleukin-1 beta (IL-1 beta) produces physiological responses that require central neural integration and because the sympathetic nervous system mediates physiological responses to environmental stress, we hypothesized that IL-1 beta modulates SND responses to acute physical stress. Therefore, this study examined the effects of IL-1 beta (290 ng/kg iv) and mild hypothermia on renal and interscapular brown adipose tissue (IBAT) SND regulation in chloralose-anesthetized rats. IBAT SND did not change after IL-1 beta administration but was significantly increased during acute mild hypothermia, which was induced 60 min after IL-1 beta treatment. Renal SND was unchanged after IL-1 beta administration and during hypothermia. Acute hypothermia, without prior IL-1 beta administration, did not alter IBAT and renal SND. Increases in IBAT SND during sustained (120 min) hypothermia were significantly higher in IL-1 beta-treated rats compared with saline-treated rats, whereas renal SND responses to sustained hypothermia did not differ among groups. Exposure to acute cold stress after sustained hypothermia produced greater increases in IBAT SND in IL-1 beta-treated rats compared with saline-treated controls. These data suggest that IL-1 beta alters IBAT SND responses to acute and sustained hypothermia.