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Am J Physiol Heart Circ Physiol 284: H1710-H1720, 2003. First published January 9, 2003; doi:10.1152/ajpheart.00673.2002
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Vol. 284, Issue 5, H1710-H1720, May 2003

Role of paraventricular nucleus in regulation of sympathetic nerve frequency components

Michael J. Kenney, Mark L. Weiss, Tammy Mendes, Yan Wang, and Richard J. Fels

Department of Anatomy and Physiology, Kansas State University, Manhattan, Kansas 66506


    ABSTRACT
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ABSTRACT
INTRODUCTION
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RESULTS
DISCUSSION
REFERENCES

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) (GABAA receptor antagonist), but not single PVN injections of NMDA (excitatory amino acid) or NG-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.

coherence function; autospectral analysis; Sprague-Dawley rats; sympathetic nerve discharge


    INTRODUCTION
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ABSTRACT
INTRODUCTION
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DISCUSSION
REFERENCES

THE PARAVENTRICULAR NUCLEUS (PVN) of the hypothalamus is an important central nervous system site for autonomic and neuroendocrine regulation (4, 22, 32, 33). The PVN contains a complex profile of excitatory and inhibitory neurotransmitters and neuromodulators (33). With respect to sympathetic nerve discharge (SND) regulation, PVN glutamate, nitric oxide (NO), and gamma -aminobutyric acid (GABA) have received considerable attention.

Ionotropic glutamate receptor subunit mRNAs are expressed in the PVN (10) and glutamate administration into the PVN of conscious rats increases heart rate (HR) (24), arterial pressure (AP) (12, 24), renal SND (12), and plasma norepinephrine concentrations (24). In addition, PVN microinjection of the excitatory amino acid, N-methyl-D-aspartic acid (NMDA), increases renal SND and AP in anesthetized rats (21), suggesting that activation of PVN NMDA receptors is excitatory to SND.

The results of several studies (11, 37) suggest that PVN NO is inhibitory to SND. PVN microinjection of nitroprusside, an NO donor, decreases AP (11, 37, 38), HR (37, 38), and renal SND (37, 38), whereas PVN administration of the nitric oxide synthase inhibitor, NG-monomethyl-L-arginine (L-NMMA), increases AP, HR, and renal SND (37). Activation of the NMDA receptor in the PVN releases NO, which inhibits NMDA-mediated increases in SND (21), suggesting interactions between excitatory and inhibitory PVN neurotransmitters in SND regulation.

Concerning GABA, several lines of evidence suggest that a PVN GABAergic system tonically inhibits SND (25, 26). PVN microinjection of muscimol, a GABA receptor agonist, decreases AP and renal SND (38). In contrast, PVN administration of bicuculline (BIC), a GABAA receptor antagonist, increases AP (17, 25, 26, 34), HR (17, 25, 26, 34), renal SND (17, 34, 38), and splenic SND (17). Zhang and Patel (38) reported that GABA mediates the renal sympathoinhibitory effect of PVN NO, suggesting interactions between PVN inhibitory neurotransmitters. Important relative to the current study, PVN BIC microinjection transforms the cardiac-related SND bursting pattern in baroreceptor-innervated rats to one characterized by the presence of low-frequency bursts (17), providing evidence that PVN GABAA receptor antagonism alters the SND bursting pattern. However, the role of the PVN in the regulation of SND frequency components is poorly understood because, with the exception of GABA, little is known about the effect of other PVN neurotransmitters on SND frequency components.

The aims of the present study were to examine (using autospectral and coherence analyses) the role of and interactions between PVN NO, GABA, and the NMDA-glutamate receptor in regulation of SND frequency components and to determine whether PVN BIC-induced changes in the SND bursting pattern alter peripheral blood flow. Experiments were completed to address four questions. First, are changes in SND frequency components unique to the disinhibition of PVN neurons produced by BIC microinjection or are SND frequency components altered after PVN microinjection of the excitatory amino acid NMDA? Second, does antagonism of PVN NO synthase (produced by L-NMMA microinjection) alter SND frequency components similar to the disinhibition of PVN neurons produced by BIC microinjection? Third, does PVN regulation of SND frequency components involve interactions between different neurotransmitters? To address this question, we determined the effect of combined PVN microinjections of L-NMMA and NMDA on SND frequency components and determined the effect of PVN microinjection of kynurenic acid (KYN; excitatory amino acid receptor antagonist) on PVN BIC-induced changes in SND frequency components. Fourth, what is the functional significance of PVN disinhibition produced by BIC administration? To address this question, renal and splenic blood flow responses to PVN BIC microinjection were measured.

Why study the central neural regulation of SND frequency components? Efferent sympathetic nerve outflow is characterized by the presence of synchronized discharge bursts that demonstrate several discharge patterns, including cardiac- and respiratory-related oscillations (1-3, 7, 8, 14, 16, 39). Importantly, the pattern of electrical stimulation of sympathetic nerves can influence renal function (6), neurotransmitter release (30), and vascular responses (28). The SND bursting pattern can be altered during various experimental interventions, including hyperthermia (15) and hypothermia (16). Alterations in the SND bursting pattern during elevations in internal body temperature contribute to hyperthermia-induced increases in the level of sympathetic nerve activity (15), suggesting that pattern transformation is a strategy used for mediating sympathoexcitation to acute heat stress. Therefore, studies designed to elucidate PVN mechanisms regulating SND frequency components are important because the SND bursting pattern is involved in physiological regulation at rest and in response to acute physical stress, and because the PVN is considered an integrative center for autonomic and neuroendocrine regulation (33, 36).


    METHODS
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The Institutional Animal Care and Use Committee approved the surgical procedures and experimental protocols.

General procedures. Male Sprague-Dawley rats (300-350 g) were anesthetized with 50-60 mg/kg ip methohexital sodium, followed by 50 mg/kg iv alpha -chloralose (initial dose), artificially ventilated, and paralyzed with 5-10 mg/kg iv gallamine triethiodide (initial dose). Catheters were placed in the femoral vein for drug administration, including maintenance doses of alpha -chloralose (35-45 mg · kg-1 · h-1), methohexital sodium (10-20 mg/kg before and during surgical interventions), and gallamine triethiodide (10-15 mg · kg-1 · h-1). End-tidal CO2 was kept near 4.5% by adjusting the frequency of ventilation. Colonic temperature was kept at 38.0°C during surgery with the use of a temperature-controlled table. Femoral arterial blood pressure and HR were recorded using standard procedures.

Bilateral denervation of the aortic arch was completed by cutting the superior laryngeal nerve near its junction with the vagus nerve and by removing the superior cervical ganglion (20). Bilateral carotid sinus denervation was completed by removing the adventitia from the carotid sinus bifurcation and applying 10% phenol to this area (20). Sinoaortic denervation (SAD) was considered complete by 1) the loss of coherence between the arterial pulse and SND and/or 2) the absence of a reflex change in SND from control levels during increases in mean arterial pressure (MAP) produced by phenylephrine hydrochloride (5 µg/kg iv) and during decreases in MAP produced by sodium nitroprusside (5 µg/kg iv).

Neural recordings. Activity was recorded biphasically with a platinum bipolar electrode after preamplification (bandpass 30-3,000 Hz) from the central end of cut renal and splenic sympathetic nerves (with the exception of experiments included in protocol IV, in which renal and splenic nerve recordings were completed from intact nerves because of the simultaneous measurement of renal and splenic blood flows). Renal and splenic nerves were isolated after a lateral incision. Sympathetic nerve potentials were full-wave rectified and integrated (time constant 10 ms), which produced a smooth tracing of the synchronized discharges (15-17). The level of activity in sympathetic nerves was quantified after integration as volts times seconds and corrected for background noise after ganglionic blockade (15 mg/kg trimethaphan camsylate) (15-17).

Central nervous system microinjections. Microinjections were made (using multibarrel glass micropipettes) unilaterally into the PVN (target site; 1.8 mm caudal to bregma, 0.7 mm lateral to the midline, 7.7 mm ventral from the surface of the brain). NMDA (100 pmol, 40 nl), L-NMMA (100 pmol, 40 nl), L-NMMA + NMDA (same doses as before), BIC (100-500 pmol, 40 nl), KYN (40 mM, 40 ml), and saline (40 nl, vehicle control) microinjections were completed in the PVN. Sites of microinjection were identified with reference to the location of the tip of the micropipette.

Recordings of blood flow velocity. Segments of the splenic and renal arteries were isolated, and a Doppler flow probe filled with ultrasonic transmission gel was placed around each vessel for measurement of blood flow velocity (19). Flow probe wires were connected to a Doppler flowmeter (19). Blood flow velocity, quantified as kilohertz Doppler shift, is directly proportional to absolute blood flow; therefore, this technique provides a relative measure of changes in blood flow (9, 19). Vascular resistance was calculated by dividing MAP by the velocity signal (19).

Brain histology. At the end of each experiment, rats received an overdose of methohexital sodium (150 mg/kg iv) and were transcardially perfused with 0.15 M NaCl (containing 3 IU/ml heparin), followed by a fixative solution consisting of 10% buffered neutral formalin (pH 7.4). The brains were removed, blocked, postfixed in buffered neutral formalin for at least 2 h, and placed in 20% sucrose for cyroprotection. Brains were frozen-sectioned at 40 µm in the coronal plane, collected into phosphate-buffered saline, and mounted on slides in serial sequence. The sections were rinsed in distilled water, air dried, cleared in xylenes, and coverslipped. The center of the injection site was localized by observing discrete clusters of latex microspheres. Microspheres could be observed in brightfield or epifluoresence (rhodamine filter cube: BP 515-560 excitation filter).

Data and statistical analysis. Autospectra and coherence analysis of the arterial pulse and SND were computed with the use of the methods and programs as previously described (14, 18). Fast Fourier transform was typically performed on 12-18 contiguous windows of data that were 5 s in duration. Autospectra and coherence functions were computed over a frequency band of 0-15 Hz. Spectral analyses provide the following information (18). The autospectrum of a signal shows the relative power present at each frequency. The coherence function (normalized cross spectrum) provides a measure of the strength of linear correlation of two signals as a function of frequency. The squared coherence value (referred to as coherence value) is 1.0 in the case of a linear system undisturbed by noise, and zero if the two signals are completely unrelated. Peak coherence values in the 0-3 Hz frequency band and at the frequency of HR [cardiac frequency (CF)] were quantified. Values in the text, tables, and figures are means ± SE. Control values of SND were taken as 0%. Statistical analyses included Student's t-test for pairwise comparisons and repeated-measures analysis of variance. P < 0.05 indicated statistical significance.


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REFERENCES

Figure 1 shows microinjection sites in the PVN and vicinity (as referenced to the tip of the micropipette) in a sagittal section modified from Paxinos and Watson (29) for experiments from each of the four protocols.


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  		Fig. 1.   Distribution of microinjection sites in the paraventricular nucleus (PVN) and vicinity. The most ventral portion of the injection tract is represented in a sagittal section modified from the one shown in the atlas of Paxinos and Watson (29). ac, Anterior commissure; AH, anterior hypothalamic area; DA, dorsal hypothalamic area; DMC, dorsal medial hypothalamic nucleus, compact part; DMD, dorsal medial hypothalamic nucleus, diffuse part; Do, dorsal hypothalamic area; LA, lateroanterior hypothalamic nucleus; oc, optic chiasm; PH, posterior hypothalamic area; SCh, suprachiasmatic nucleus; StHy, striohypothalamic nucleus; VMH, ventromedial hypothalamic nucleus.

Protocol I: effect of PVN NMDA, L-NMMA, and BIC microinjections on SND frequency components. PVN microinjection of NMDA (100 pmol, 40 nl) was completed in five experiments. Figure 2A shows traces of SND bursts (renal and splenic) and pulsatile AP recorded before and after PVN NMDA microinjection from a representative experiment. The renal and splenic SND bursting patterns remained unchanged from control after PVN NMDA microinjection. Figure 3 shows SND autospectra (top) and coherence functions (bottom) constructed during control and at 7 min after PVN NMDA microinjection. The shape and contour of the SND autospectra and coherence functions were similar before (control) and after PVN NMDA microinjection. Mean peak coherence values relating renal-splenic discharges at the CF (control, 0.87 ± 0.03; 4 ± 1 min postinjection, 0.92 ± 0.02; 7 ± 1 min postinjection, 0.92 ± 0.01) and in the 0-3 Hz frequency band (control, 0.76 ± 0.04; 4 ± 1 min postinjection, 0.80 ± 0.03; 7 ± 1 min postinjection, 0.78 ± 0.04) were unchanged from control after PVN NMDA microinjection. Peak coherence values were derived when PVN NMDA-induced sympathoexcitatory responses were present (4-7 min after PVN NMDA microinjection). Peak coherence values relating renal and splenic SND bursts were unchanged from control for up to 30 min after PVN NMDA microinjection. Levels of renal and splenic SND were significantly increased from control (renal, 28 ± 8%, 4-7 min after injection; splenic, 15 ± 3%, 4-7 min after injection), whereas MAP (control, 106 ± 6 mmHg; 4-7 min after injection, 106 ± 7 mmHg) remained unchanged after PVN NMDA microinjection (n = 5).


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  		Fig. 2.   Traces of integrated sympathetic nerve discharge (SND) bursts (renal and splenic) and pulsatile arterial pressure (AP) recorded before (control) and after PVN microinjections of 100 pmol of N-methyl-D-aspartic acid (NMDA; A), 100 pmol of NG-monomethyl-L-arginine (L-NMMA; B), and 100 pmol of bicuculline (BIC; C). Horizontal calibration is 500 ms.



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  		Fig. 3.   Frequency-domain relationships (from a representative experiment) between renal and splenic (Ren/Spl) SND bursts before (control) and after (7 min) microinjection of NMDA (100 pmol) into the PVN. The two top panels for each period are individual autospectra, and bottom panels are nerve-to-nerve coherence functions. Frequency band is displayed from 0 to 15 Hz. SND total power was increased after PVN NMDA microinjection (renal SND, 43%; splenic SND, 24%).

PVN microinjection of L-NMMA (100 pmol, 40 nl) was completed in five experiments. Renal and splenic SND bursting patterns (Fig. 2B shows original traces from a representative experiment) and mean peak coherence values relating renal-splenic discharges at the CF (control, 0.89 ± 0.02; 8 ± 1 min postinjection, 0.88 ± 0.02; 11 ± 1 min postinjection, 0.89 ± 0.02) and in the 0-3 Hz frequency band (control, 0.74 ± 0.08; 8 ± 1 min postinjection, 0.81 ± 0.04; 11 ± 1 min postinjection, 0.79 ± 0.05) remained unchanged from control after PVN L-NMMA microinjection. Peak coherence values were derived when sympathoexcitatory responses to PVN L-NMMA were present (8-11 min after PVN L-NMMA microinjection). Peak coherence values relating renal and splenic SND bursts were unchanged from control for up to 30 min after PVN L-NMMA microinjection. Levels of renal and splenic SND were significantly increased from control (renal, 20 ± 5%, 8-11 min after injection; splenic, 11 ± 3%, 8-11 min after injection) after PVN L-NMMA microinjection whereas MAP (control, 108 ± 3 mmHg; 11 min after injection, 111 ± 3 mmHg) remained unchanged (n = 5).

PVN microinjection of BIC (200-500 pmol, 40 nl) was completed in seven experiments. BIC microinjections were completed after NMDA and L-NMMA microinjections. In contrast to sympathetic recordings completed after PVN NMDA (Fig. 2A) or L-NMMA (Fig. 2B) microinjections, SND was characterized by the presence of low-frequency bursts after PVN BIC microinjection (Fig. 2C). Consistent with our previous study (17), mean peak coherence values relating renal-splenic discharges at the CF (control, 0.88 ± 0.03; 5 min after injection, 0.55 ± 0.03, P < 0.05) were significantly reduced despite increased (15 ± 2 mmHg, 5 min after injection) MAP, whereas the coupling of discharges in the 0-3 Hz frequency band (control, 0.85 ± 0.02; 5 min after injection, 0.97 ± 0.01, P < 0.05) was significantly increased after PVN BIC microinjection. Peak increases (renal SND, 35 ± 2% from control; splenic SND, 24 ± 3% from control) in the level of SND after PVN BIC microinjection were determined in three experiments and occurred 10-15 min after PVN BIC microinjection.

Peak coherence values relating renal-splenic discharges in the 0-3 Hz frequency band (control, 0.72 ± 0.07; 5 min after injection, 0.74 ± 0.03) and at the CF (control, 0.86 ± 0.03; 5 min after injection, 0.91 ± 0.02), the level of renal and splenic nerve activity (+2 ± 2%, 5 min after injection), and MAP (control, 98 ± 2 mmHg; 5 after injection, 99 ± 2 mmHg) remained unchanged from control after PVN saline microinjection (n = 4).

Protocol II: effect of PVN L-NMMA + NMDA microinjections on SND frequency components. Combined PVN microinjections of L-NMMA and NMDA (L-NMMA microinjection completed 2 min before NMDA microinjection, L-NMMA + NMDA) were completed in 10 experiments. The SND bursting pattern, recorded at 6 ± 2 min after PVN L-NMMA + NMDA microinjections, was characterized by clusters of bursts interspersed with periods of quiescence (see Fig. 4, middle). This distinctive SND bursting pattern, which persisted for 5 ± 2 min after combined L-NMMA + NMDA microinjections, was not observed in experiments in which single PVN injections of either NMDA or L-NMMA were completed and was not similar to PVN BIC-induced changes in the SND bursting pattern [Fig. 4, compare SND traces shown in the middle (PVN L-NMMA + NMDA) and right (PVN BIC) panels]. The primary peak in the 0-3 Hz frequency band was shifted from 1.2 ± 0.3 Hz during control to 0.3 ± 0.07 Hz after PVN L-NMMA + NMDA microinjection, and the peak coherence value relating renal-splenic discharges in this frequency band was significantly increased from 0.77 ± 0.03 at 1.2 ± 0.3 Hz during control to 0.90 ± 0.03 at 0.3 ± 0.07 Hz after PVN L-NMMA + NMDA. The peak coherence value relating renal-splenic discharges at the CF (control, 0.84 ± 0.03; L-NMMA + NMDA, 0.91 ± 0.03) remained unchanged after PVN microinjection of L-NMMA + NMDA. Renal and splenic SND were significantly increased from control (renal, 43 ± 15%, 7 ± 3 min after injection; splenic, 21 ± 3%, 7 ± 3 min after injection), whereas MAP remained unchanged (control, 113 ± 6 mmHg; 7 ± 3 min, 112 ± 5 mmHg) after PVN L-NMMA + NMDA microinjections.


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  		Fig. 4.   Traces of SND bursts (renal and splenic) and AP recorded from a representative experiment during control, 12 min after microinjection of L-NMMA (100 pmol) and NMDA (100 pmol) into the PVN (PVN L-NMMA + NMDA), and 3 min after PVN microinjection of BIC (200 pmol). The PVN BIC microinjection was completed 30 min after the combined injection of L-NMMA + NMDA. Horizontal calibration is 500 ms.

The alternating periods of SND bursts and quiescence observed after PVN L-NMMA + NMDA microinjections were coupled to a slow fluctuation (0.3-0.4 Hz) in arterial blood pressure; specifically, cardiac-related bursts were present when arterial pressure declined, whereas periods of quiescence were present when arterial pressure increased (see Fig. 5A). Peak-to-trough changes in arterial blood pressure observed during slow fluctuations in arterial pressure ranged from 3 to 6 mmHg for systolic blood pressure and from 2 to 4 mmHg for diastolic blood pressure (n = 4). After PVN L-NMMA + NMDA microinjection in SAD rats (n = 4), renal and splenic SND recordings were characterized by the presence of noncardiac-related bursts and were devoid of periods of SND quiescence (see Fig. 5B).


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  		Fig. 5.   Traces of SND bursts (renal and splenic) and AP recorded after microinjection of L-NMMA and NMDA (100 pmol of each drug) into the PVN in a rat with intact arterial baroreceptors (A) and in a sinoaortic denervated (SAD) rat (B). Horizontal calibration is 500 ms.

Protocol III: effect of KYN + BIC microinjections on SND frequency components. PVN BIC-induced changes in the SND bursting pattern were prevented by PVN pretreatment with KYN (40 mM, 40 nl) in three experiments. PVN KYN microinjections were completed 2-3 min before PVN BIC microinjections. Figure 6 shows the results of frequency-domain analyses of renal and splenic SND from one representative experiment. SND autospectra (Fig. 6, top) and coherence functions (Fig. 6, bottom) constructed 2 min after PVN KYN microinjection were similar to those constructed during control. PVN BIC microinjection was completed 3 min after PVN KYN microinjection. SND autospectra and coherence functions constructed after (3 min) PVN BIC microinjection were similar to those constructed during control and after PVN KYN. Mean peak coherence values in the 0-3 Hz frequency band (control, 0.90 ± 0.03; 2 min after PVN KYN, 0.93 ± 0.01; 3-5 min after PVN BIC, 0.88 ± 0.02) and at the CF (control, 0.93 ± 0.01; 2 min after PVN KYN, 0.91 ± 0.01; 3-5 min after PVN BIC, 0.92 ± 0.01) remained unchanged from control values after PVN pretreatment with KYN and after PVN BIC following PVN KYN microinjection. In contrast, PVN BIC microinjection (100 pmol, 40 nl, n = 3) 30 min after PVN KYN microinjection and PVN BIC microinjection (100 pmol, 40 nl) 2 min after PVN pretreatment with saline (40 nl, n = 2) changed the SND bursting pattern similar to that shown in Fig. 2C and Fig. 8 (PVN BIC).


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  		Fig. 6.   Frequency-domain relationships (from a representative experiment) between renal and splenic sympathetic nerve discharge bursts during control, 2 min after microinjection of kynurenic acid (KYN; 40 mM) into the PVN, and 3 min after PVN microinjection of BIC (100 pmol). PVN BIC microinjection was completed 3 min after the PVN KYN microinjection. Top two panels during each period are individual SND autospectra and the bottom panels are nerve-to-nerve coherence functions. Frequency band is displayed from 0 to 15 Hz. SND total power was not markedly altered after PVN KYN microinjection (renal SND, -4%; splenic SND, -3%) or after PVN BIC microinjection after PVN KYN microinjection (renal SND, +10%; splenic SND, +2%).

PVN BIC-induced changes in the SND bursting pattern were reversed by PVN KYN microinjection in four experiments. Figure 7 shows traces of renal and splenic SND bursts during control and after consecutive PVN microinjections of BIC (100 pmol, 40 nl), saline (40 nl), and KYN (40 mM, 40 nl) from one representative experiment. As expected, PVN BIC microinjection altered the SND bursting pattern and the low-frequency bursts persisted after PVN saline microinjection (traces are from 15-17 and 45-47 s after saline microinjection). In contrast, after PVN KYN microinjection (PVN KYN after PVN BIC, traces are from 15-17 and 45-47 s after KYN microinjection), low-frequency SND bursts were eliminated, and SND traces were characterized by the presence of cardiac-related bursts. The results of autospectral (Fig. 8, top) and coherence (Fig. 8, bottom) analyses of renal and splenic SND from one representative experiment are shown in Fig. 8. Cardiac-related peaks in the SND autospectra were eliminated and low-frequency peaks were evident in SND autospectra after PVN BIC microinjection. SND autospectra (6 contiguous windows of data that were 5 s in duration) initiated at 10 and 60 s after PVN KYN microinjection contained peaks at the frequency of the cardiac cycle, although the cardiac-related peaks were more prominent in the renal SND autospectra.


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  		Fig. 7.   Traces of integrated SND bursts (renal and splenic) and AP recorded during control, 3 min after PVN microinjection of BIC (100 pmol) (PVN BIC), 15 and 45 s after PVN saline microinjection (PVN saline after PVN BIC), and 15 and 45 s after PVN KYN microinjection (PVN KYN after PVN BIC). The PVN saline microinjection was completed during the third minute after PVN BIC microinjection, whereas the PVN KYN microinjection was completed during the fourth minute after PVN BIC microinjection. Horizontal calibration is 500 ms.



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  		Fig. 8.   Frequency-domain relationships (from a representative experiment) between renal and splenic SND bursts during; control, 3 min after PVN BIC (100 pmol) microinjection, 10 s after PVN KYN (40 mM) microinjection (PVN KYN after BIC, 10 s), and 60 s after PVN KYN microinjection (PVN KYN after BIC, 60 s). The KYN microinjection was completed 3 min after PVN BIC microinjection. The two top panels for each period are individual autospectra and the bottom panels are nerve-to-nerve coherence functions. Frequency band is displayed from 0 to 15 Hz. SND total power was reduced after PVN BIC microinjection (renal SND, -14%; splenic SND, -5%), increased during the postinjection period initiated 10 s after PVN KYN after PVN BIC (renal SND, +13%; splenic SND, +12%), and relatively unchanged during the postinjection period initiated 60 s after PVN KYN after PVN BIC (renal SND, +2%; splenic SND, -5%).

Mean peak coherence values relating renal-splenic discharges at the CF were significantly decreased, whereas those relating discharges in the 0-3 Hz frequency band were significantly increased after PVN BIC microinjection (Table 1). Mean peak coherence values derived from coherence functions constructed 30-60 s after PVN KYN microinjection were not different from control for the CF but remained slightly but significantly increased for discharges in the 0-3 Hz frequency band (Table 1). Coherence values relating renal-splenic discharges at the CF and in the 0-3 Hz frequency band were significantly changed from control after PVN BIC and remained this way after PVN saline microinjection (1-2 min after PVN BIC, n = 4).

                              
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  		Table 1.   Peak coherence values relating discharges in renal-splenic sympathetic nerve pairs

Protocol IV: splenic and renal blood flow responses to PVN BIC microinjection. Splenic and renal blood flow responses to PVN microinjection of BIC (100 pmol, 40 nl) were determined in four experiments. Three minutes after PVN BIC microinjections, low-frequency bursts were present in splenic and renal SND recordings (similar to those shown in Figs. 2, 4, and 7), the level of renal and splenic sympathetic nerve activity was not significantly changed from control (+7 ± 13%, renal and splenic SND combined), blood flow in the splenic and renal arteries tended to be decreased (-16 ± 9%) but was not significantly decreased from control, and MAP was increased (15 ± 2 mmHg). Because of BIC-induced increases in MAP, renal and splenic blood flows were normalized to MAP and expressed as resistance. Vascular resistance was significantly increased 37 ± 16% (splenic and renal data combined, P < 0.05) 3 min after PVN BIC microinjection.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The current results provide four new findings concerning the role of the PVN in regulation of SND frequency components in chloralose-anesthetized rats. First, in contrast to PVN BIC microinjection, single PVN microinjections of NMDA or L-NMMA did not alter SND frequency components. Second, combined PVN L-NMMA and NMDA microinjections altered the SND bursting pattern; however, the observed pattern change was unlike that produced by PVN GABAA receptor antagonism. Third, blockade of excitatory amino acid receptors in the PVN with KYN prevented and reversed PVN BIC-induced changes in the SND bursting pattern. Fourth, PVN BIC microinjection increased renal and splenic vascular resistance despite the fact that the total level of activity in these nerves was not changed. It should be noted that we cannot exclude the possibility that the perinuclear region of the PVN may play a role in mediating the observed SND responses because some injection sites were just outside the boundaries of the PVN.

It is known that a PVN GABAergic system exerts a tonic inhibitory effect on the level of efferent sympathetic nerve activity (25, 26) and that PVN GABAA receptor antagonism alters SND frequency components (Ref. 17 and current study). However, the neurotransmitter profile of the PVN is complex (33), and the role of other PVN neurotransmitters, including glutamate and NO, in regulation of SND frequency components is not well established. Several lines of evidence suggest an important functional role of PVN glutamate in SND regulation. As demonstrated by Herman et al. (10), ionotropic glutamate receptor subunit mRNAs for NMDA-, alpha -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-, and kainate-preferring glutamate receptors are expressed in the PVN. In addition, the level of activity in efferent sympathetic nerves is altered after PVN microinjection of glutamate (12, 13), kainic acid (31), D,L-homocysteic acid (5, 23), and NMDA (21).

Does activation of PVN NMDA excitatory amino acid receptors affect SND frequency components? The current results suggest this is not the case because renal and splenic SND autospectra and renal-splenic coherence functions remained unchanged from control after PVN NMDA microinjection, despite activation of sympathetic neural circuits as demonstrated by significant increases in the level of renal and splenic SND after PVN NMDA microinjection. In contrast to the findings of Li et al. (21), in the present study, MAP remained unchanged after PVN NMDA microinjection despite increased renal and splenic SND.

Several lines of evidence suggest that NO influences PVN neuronal activity. For example, NO synthase positive neurons are located in the PVN (27, 35), PVN microinjection of sodium nitroprusside (NO donor) reduces the level of renal sympathetic nerve activity (37, 38), and PVN microinjection of NO synthase inhibitors increases renal SND (37). These data suggest that in addition to GABA, PVN NO provides tonic inhibition to sympathetic nerve activity. In fact, Zhang and Patel (38) reported that the inhibitory influence of PVN NO on the level of efferent renal SND is mediated by GABA. On the basis of these results, we hypothesized that PVN NO synthase inhibition would alter the SND bursting pattern and the coupling of SND bursts similar to that produced by PVN microinjection of bicuculline (17). This was not the case, however, because renal and splenic SND autospectra and renal-splenic coherence functions remained unchanged after PVN L-NMMA microinjection, despite increased renal and splenic SND after PVN L-NMMA microinjection. In contrast to the findings of Zhang et al. (37), in the present study, MAP remained unchanged after PVN L-NMMA microinjection, despite increased renal and splenic SND.

As reported by Li et al. (21), PVN NMDA-induced renal sympathoexcitation is augmented by inhibition of endogenous NO synthesis by PVN L-NMMA microinjection, demonstrating functional interactions between PVN NO and at least one type of glutamate receptor. On the basis of this finding, we hypothesized that activation of PVN NMDA excitatory amino acid receptors after inhibition of endogenous NO would alter SND frequency components. The current results demonstrate this is the case; however, the SND pattern change after PVN microinjection of L-NMMA + NMDA is unlike that produced by PVN BIC microinjection in two important ways. First, cardiac-related SND bursts are virtually eliminated after PVN BIC microinjection in baroreceptor-innervated rats (Ref. 17 and current study), whereas they persist after combined PVN microinjections of L-NMMA + NMDA, although they are interspersed with periods of quiescence. Second, PVN BIC-induced changes in the SND bursting pattern occur in baroreceptor-innervated and -denervated rats (17), whereas the SND bursting pattern to PVN L-NMMA + NMDA microinjections is not observed in SAD rats, suggesting an important role for the arterial baroreceptors in mediating SND pattern changes observed after combined PVN nitric oxide synthase inhibition and NMDA receptor activation. The prolonged episodes of SND inhibition observed after combined PVN microinjections of L-NMMA + NMDA were evident despite small and slowly developing increases in arterial pressure, suggesting sensitization of the arterial baroreflex regulation of efferent sympathetic nerve outflow. The importance of functional interactions between different PVN neurotransmitters in regulation of SND frequency components is further supported by the finding that blockade of ionotropic glutamate receptors in the PVN prevented and reversed SND frequency-domain changes evoked by blockade of PVN GABAA receptors.

What is the functional significance of the PVN BIC-induced SND pattern change? In the present study, despite significant increases in perfusion pressure, renal and splenic blood flow remained unchanged from control after PVN BIC microinjection due to significant increases in calculated vascular resistance. The increase in resistance was evident despite the fact that the total level of activity in the renal and splenic nerves remained unchanged during the first few minutes after PVN BIC microinjection, suggesting an important functional role for transformation of the SND pattern to low-frequency bursts. The results of other studies support a role for the SND bursting pattern in physiological regulation. For example, the pattern of electrical stimulation of efferent sympathetic nerves affects physiological responses in the kidney (vasoconstriction and urinary sodium excretion) (6), neurotransmitter release in the pig spleen (30), and contractile responses of rat mesenteric arteries (28). In addition, alterations in the SND bursting pattern during heating contribute to hyperthermia-induced sympathoexcitation (15).

Regulation of processes essential for maintaining physiological homeostasis at rest and for mediating responses to acute physical stress requires the generation of complex output profiles by central sympathetic neural circuits. As stated previously, changing the pattern of SND bursts is one strategy used by sympathetic neural circuits to alter efferent SND. The present findings 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 regulation of the SND bursting pattern.


    ACKNOWLEDGEMENTS

This study was supported by National Heart, Lung, and Blood Institute Grants HL-65346 and HL-69755 (to M. J. Kenney).


    FOOTNOTES

Address for reprint requests and other correspondence: M. J. Kenney, Dept. Anatomy and Physiology, Coles Hall 228, Kansas State Univ., 1600 Denison Ave., Manhattan, KS 66506 (E-mail: Kenny{at}vet.ksu.edu).

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. Section 1734 solely to indicate this fact.

First published January 9, 2003;10.1152/ajpheart.00673.2002

Received 31 July 2002; accepted in final form 6 January 2003.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Barman, SM. Brainstem control of cardiovascular function. In: Brainstem Mechanisms of Behavior, edited by Klemm WR, and Vertes RP.. New York: Wiley, 1990, p. 353-381.

2.   Barman, SM, and Gebber GL. Sympathetic nerve rhythm of brain stem origin. Am J Physiol Regul Integr Comp Physiol 239: R42-R47, 1980[Free Full Text].

3.   Claassen, DE, Fels RJ, and Kenney MJ. Altered frequency characteristics of sympathetic nerve activity after sustained elevation in arterial pressure. Am J Physiol Regul Integr Comp Physiol 274: R694-R703, 1998[Abstract/Free Full Text].

4.   Dampney, RAL Functional organization of central pathways regulating the cardiovascular system. Physiol Rev 74: 323-364, 1994[Free Full Text].

5.   Deering, J, and Coote JH. Paraventricular neurons elicit a volume expansion-like change of activity in sympathetic nerves to the heart and kidney in the rabbit. Experimental Physiology 85: 177-186, 2000[Abstract].

6.   DiBona, GF, and Sawin LL. Functional significance of the pattern of renal sympathetic nerve activation. Am J Physiol Regul Integr Comp Physiol 277: R346-R353, 1999[Abstract/Free Full Text].

7.   Gebber, GL. Central oscillators responsible for sympathetic nerve discharge. Am J Physiol Heart Circ Physiol 239: H143-H155, 1980[Abstract/Free Full Text].

8.   Gebber, GL. Central determinants of sympathetic nerve discharge. In: Central Regulation of Autonomic Functions, edited by Loewy AD, and Spyer KM.. New York: Oxford University Press, 1990, p. 126-144.

9.   Haywood, JR, Shaffer RA, Fastenow C, Fink GD, and Brody MJ. Regional blood flow measurement with pulsed Doppler flowmeter in conscious rat. Am J Physiol Heart Circ Physiol 241: H273-H278, 1981[Abstract/Free Full Text].

10.   Herman, JP, Eyigor O, Ziegler DR, and Jennes L. Expression of ionotropic glutamate receptor subunit mRNAs in the hypothalamic paraventricular nucleus of the rat. J Comp Neurol 422: 352-362, 2000[Web of Science][Medline].

11.   Horn, T, Smith PM, McLaughlin BE, Bauce L, Marks GS, Pittman QJ, and Ferguson AV. Nitric oxide actions in paraventricular nucleus: cardiovascular and neurochemical implications. Am J Physiol Regul Integr Comp Physiol 266: R306-R313, 1994[Abstract/Free Full Text].

12.   Kannan, H, Hayashida Y, and Yamashita H. Increase in sympathetic outflow by paraventricular nucleus stimulation in awake rats. Am J Physiol Regul Integr Comp Physiol 256: R1325-R1330, 1989[Abstract/Free Full Text].

13.   Katafuchi, T, Oomura Y, and Kurosawa M. Effects of chemical stimulation of paraventricular nucleus on adrenal and renal nerve activity in rats. Neurosci Lett 86: 195-200, 1988[Web of Science][Medline].

14.   Kenney, MJ. Frequency characteristics of sympathetic nerve discharge in anesthetized rats. Am J Physiol Regul Integr Comp Physiol 267: R830-R840, 1994[Abstract/Free Full Text].

15.   Kenney, MJ, Claassen DE, Bishop MR, and Fels RJ. Regulation of the sympathetic nerve discharge bursting pattern during heat stress. Am J Physiol Regul Integr Comp Physiol 275: R1992-R2001, 1998[Abstract/Free Full Text].

16.   Kenney, MJ, Claassen DE, Fels RJ, and Saindon CS. Cold stress alters characteristics of sympathetic nerve discharge bursts. J Appl Physiol 87: 732-742, 1999[Abstract/Free Full Text].

17.   Kenney, MJ, Weiss ML, Patel KP, Wang Y, and Fels RJ. Paraventricular nucleus bicuculline alters frequency components of sympathetic nerve discharge bursts. Am J Physiol Heart Circ Physiol 281: H1233-H1241, 2001[Abstract/Free Full Text].

18.   Kocsis, B, Gebber GL, Barman SM, and Kenney MJ. Relationships between activity of sympathetic nerve pairs: phase and coherence. Am J Physiol Regul Integr Comp Physiol 259: R549-R560, 1990[Abstract/Free Full Text].

19.   Kregel, KC, Kenney MJ, Massett MP, Morgan DA, and Lewis SJ. Role of nitrosyl factors in the hemodynamic adjustments to heat stress in the rat. Am J Physiol Heart Circ Physiol 273: H1537-H1543, 1997[Abstract/Free Full Text].

20.   Krieger, EM. Neurogenic hypertension in the rat. Circ Res 15: 511-521, 1964[Abstract/Free Full Text].

21.   Li, YF, Mayhan WG, and Patel KP. NMDA-mediated increase in renal sympathetic nerve discharge within the PVN: role of nitric oxide. Am J Physiol Heart Circ Physiol 281: H2328-H2336, 2001[Abstract/Free Full Text].

22.   Loewy, AD. Central autonomic pathways. In: Central Regulation of Autonomic Functions, edited by Loewy AD, and Spyer KM.. New York: Oxford University Press, 1990, p. 88-103.

23.   Lu, XZ, Sun XY, and Yao T. Inhibition of renal nerve activity induced by chemical stimulation of the paraventricular nucleus: mediation of the vasopressinergic spinally-projecting pathway. Chi J Physiol Sci 7: 215-221, 1991.

24.   Martin, DS, and Haywood JR. Sympathetic nervous system activation by glutamate injections into the paraventricular nucleus. Brain Res 577: 261-267, 1992[Web of Science][Medline].

25.   Martin, DS, and Haywood JR. Hemodynamic responses to paraventricular nucleus disinhibition with bicuculline in conscious rats. Am J Physiol Heart Circ Physiol 265: H1727-H1733, 1993[Abstract/Free Full Text].

26.   Martin, DS, Segura T, and Haywood JR. Cardiovascular responses to bicuculline in the paraventricular nucleus of the rat. Hypertension 18: 48-55, 1991[Abstract/Free Full Text].

27.   Miyagawa, A, Okamura H, and Ibata Y. Coexistence of oxytocin and NADPH-diaphorase in magnocellular neurons of the paraventricular and the supraoptic nuclei of the rat hypothalamus. Neurosci Lett 171: 13-16, 1994[Web of Science][Medline].

28.   Nilsson, H, Ljung B, Sjoblom N, and Wallin BG. The influence of the sympathetic impulse pattern on contractile responses of rat mesenteric arteries and veins. Acta Physiol Scand 123: 303-309, 1985[Web of Science][Medline].

29.   Paxinos, G, and Watson C. The Rat Brain in Stereotaxic Coordinates (3rd ed.). San Diego, CA: Academic, 1997.

30.   Pernow, J, Schwieler J, Kahan T, Hjemdahl P, Oberle J, Wallin BG, and Lundberg JM. Influence of sympathetic discharge pattern on norepinephrine and neuropeptide Y release. Am J Physiol Heart Circ Physiol 257: H866-H872, 1989[Abstract/Free Full Text].

31.   Porter, JP. Contribution of spinal N-methyl-D-aspartic acid receptors to control of sympathetic outflow by the paraventricular nucleus. Brain Res Bull 32: 653-660, 1993[Web of Science][Medline].

32.   Sun, MK. Central neural organization and control of sympathetic nervous system in mammals. Prog Neurobiology 47: 157-233, 1995[Web of Science][Medline].

33.   Swanson, LW, and Sawchenko PE. Hypothalamic integration: organization of the paraventricular and supraoptic nuclei. Ann Rev Neurosci 6: 269-324, 1983[Web of Science][Medline].

34.   Tagawa, T, and Dampney RAL AT1 receptors mediate excitatory inputs to rostral ventrolateral medulla pressor neurons from hypothalamus. Hypertension 34: 1301-1307, 1999[Abstract/Free Full Text].

35.   Vincent, SR, and Kimura H. Histochemical mapping of nitric oxide synthase in the rat brain. Neurosci 46: 755-784, 1992.

36.   Yang, H, Wang L, and Gong J. Evidence for hypothalamic paraventricular nucleus as an integrative center of neuroimmunomodulation. Neuroimmunomodulation 4: 120-127, 1997[Web of Science][Medline].

37.   Zhang, K, Mayhan WG, and Patel KP. Nitric oxide within the paraventricular nucleus mediates changes in renal sympathetic nerve activity. Am J Physiol Regul Integr Comp Physiol 273: R864-R872, 1997[Abstract/Free Full Text].

38.   Zhang, K, and Patel KP. Effect of nitric oxide within the paraventricular nucleus on renal sympathetic nerve discharge: role of GABA. Am J Physiol Regul Integr Comp Physiol 275: R728-R734, 1998[Abstract/Free Full Text].

39.   Zhong, S, Zhou SY, Gebber GL, and Barman SM. Coupled oscillators account for the slow rhythms in sympathetic nerve discharge and phrenic nerve activity. Am J Physiol Regul Integr Comp Physiol 272: R1314-R1324, 1997[Abstract/Free Full Text].

Am J Physiol Heart Circ Physiol 284(5):H1710-H1720
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