Aortic Nerve Research Articles

Integrin ligands block mechanical signal transduction in baroreceptors.

Baroreceptors are nerve endings located in the adventitia of the carotid sinus and aortic arch. They act as a mechanoelectrical transducer that can sense the tension stimulation exerted on the blood vessel wall by the rise in blood pressure and transduce the mechanical force into discharge of the nerve endings. However, the molecular identity of mechanical signal transduction from the vessel wall to the baroreceptor is not clear. We discovered that exogenous integrin ligands, such as RGD, IKVAV, YIGSR, PHSRN, and KNEED, could restrain pressure-dependent discharge of the aortic nerve in a dose-dependent and reversible manner. Perfusion of RGD at the baroreceptor site in vivo can block the baroreceptor reflex. An immunohistochemistry study showed the binding of exogenous RGD to the nerve endings under the adventitia of the rat aortic arch, which may competitively block the binding of integrins to ligand motifs in extracellular matrix. These findings suggest that connection of integrins with extracellular matrix plays an important role in the mechanical coupling process between vessel walls and arterial baroreceptors.

Abstract MP22: Dietary Salt Restriction And High Salt Intake Exaggerate Sympathetic Reflexes And Sensitize Central Autonomic Networks: Potential Mechanisms Of The J-shaped Curve

Observational cohort studies suggest that severe salt restriction increases cardiovascular morbidity/mortality, and the relationship between cardiovascular morbidity and dietary salt intake resembles a J-shaped curve. A high salt diet exaggerates sympathetic nerve activity (SNA) and arterial blood pressure (ABP) responses to several cardiovascular reflexes in salt-resistant animals. This study assessed whether salt restriction also exaggerates cardiovascular reflex responses and sensitizes central autonomic networks. To test this hypothesis, male Sprague-Dawley rats were fed low (0.01% NaCl), normal (0.1% NaCl), and high (4.0% NaCl) salt diet for 14-21 days. Baseline mean ABP was not different across groups (low: 104±4, normal: 107±4, high: 107±4mmHg). Activation of sciatic afferents (1ms pulse, 500uA, 5s duration, 2-20Hz) produced significantly greater increases in renal SNA (5Hz; low: 196±12, normal: 136±9, high: 177±8%, n=8, P<0.05) and ABP (5Hz; low: 29±3, normal: 16±1, high: 24±2 mmHg, n=8, P<0.05) of rats fed low and high versus normal NaCl diets. Activation of the aortic depressor nerve (2ms pulse, 500uA, 15s duration, 2-20Hz) produced significantly greater decreases in renal SNA (5Hz; low: -55±9, normal: -34±8, high: -63±13%, n=7-8, P<0.05) and ABP (5Hz; low: -31±3, normal: -15±5, high: -32±5 mmHg, n=7-8, P<0.05) of rats fed low and high versus normal NaCl diets. To test whether dietary salt intake sensitized central sympathetic circuits, microinjection of L-glutamate (0.1-1nmol, 30nL) in the rostral ventrolateral medulla produced significantly greater increases in renal SNA (0.1nmol; low: 212±15, normal: 149±8, high: 183±17%, n=7-8, P<0.05) and ABP (0.1Hz; low: 20±2, normal: 12±2, high: 22±2 mmHg, n=7-8, P<0.05) of rats fed low and high versus normal NaCl diets. Finally, rats fed low or high NaCl versus normal NaCl diets displayed exaggerated cardiovascular responses to cage switch or mild restraint and increased 24-h blood pressure variability. The present findings show that severe salt restriction and excess dietary salt intake exaggerate sympathetic and cardiovascular responses, and may be explained by a parallel change in the sensitivity of central autonomic networks to resemble a J-shaped curve.

ASIC2 Synergizes with TRPV1 in the Mechano-Electrical Transduction of Arterial Baroreceptors.

Mechanosensitive ion channels (MSCs) are key molecules in the mechano-electrical transduction of arterial baroreceptors. Among them, acid-sensing ion channel 2 (ASIC2) and transient receptor potential vanilloid subfamily member 1 (TRPV1) have been studied extensively and documented to play important roles. In this study, experiments using aortic arch-aortic nerve preparations isolated from rats revealed that both ASIC2 and TRPV1 are functionally necessary, as blocking either abrogated nearly all pressure-dependent neural discharge. However, whether ASIC2 and TRPV1 work in coordination remained unclear. So we carried out cell-attached patch-clamp recordings in HEK293T cells co-expressing ASIC2 and TRPV1 and found that inhibition of ASIC2 completely blocked stretch-activated currents while inhibition of TRPV1 only partially blocked these currents. Immunofluorescence staining of aortic arch-aortic adventitia from rats showed that ASIC2 and TRPV1 are co-localized in the aortic nerve endings, and co-immunoprecipitation assays confirmed that the two proteins form a compact complex in HEK293T cells and in baroreceptors. Moreover, protein modeling analysis, exogenous co-immunoprecipitation assays, and biotin pull-down assays indicated that ASIC2 and TRPV1 interact directly. In summary, our research suggests that ASIC2 and TRPV1 form a compact complex and function synergistically in the mechano-electrical transduction of arterial baroreceptors. The model of synergism between MSCs may have important biological significance beyond ASIC2 and TRPV1.

Standard Pancreatoduodenectomy Versus Radical Pancreatoduodenectomy With Retroperitoneal Nerve Resection in Patients With Pancreatic Head Cancer (RAPNER): A Multicentre Randomized Controlled Trial

Background: Pancreatic cancer is a highly fatal disease and is becoming an increasingly common cause of cancer mortality. The extent of pancreatoduodenectomy for pancreatic head cancer is still controversial, and more high-level clinical evidence is needed. We compared the outcomes of standard pancreatoduodenectomy (SPD) with those of radical pancreatoduodenectomy (RPD) with retroperitoneal nerve resection in pancreatic head cancer patients in this multicentre randomized controlled trial. Methods: RAPNER was open-label, multicentre randomised trial, patients aged 18–80 years with stage I or II pancreatic head cancer undergoing curative resection were enrolled at 6 centres from China. Patients were allocated (1:1) by use of a computer-based randomisation service to assigned to receive standard pancreatoduodenectomy (PD) or radical PD, with the latter followed by the combined dissection of additional lymph nodes (LNs), nerves and soft tissues 270° on the right side surrounding the superior mesenteric artery and celiac axis. This trial is registered with Chinese Clinical Trial Registry (http://www.chictr.org.cn), ChiCTR-TRC-12002548. Findings: Between October 3, 2012, and February 15, 2017, 468 patients were enrolled; of these, 400 were randomly assigned to the RPD group (n=201) or the SPD group (n=199). The operation time was longer with RPD than with SPD, but the R1 resection rate was slightly higher in the SPD group (12·56%) than in the RPD group (8·46%). Both the total number of retrieved LNs and number of positive LNs were significantly higher in the RPD group (20 (7-63) and 1 (0-29)) than in the SPD group (15 (7-63) and 1 (0-11)). Both the morbidity and inpatient mortality rates were comparable between the two groups. The median overall survival (OS) of the RPD and SPD groups were 23·0 months and 20·2 months (p=0·100), and the disease-free survival (DFS) were 15·6 months and 13·5 months (p=0·060). In the subgroup with a preoperative CA19-9 level less than 200 U/ml, the median OS in the RPD group was 30·8 months, which was significantly longer than that in the SPD group (20·9 months; p=0·009), and the median DFS was significantly longer in the RPD group than in the SPD group (22·3 months vs. 15·0 months, p=0·002). Furthermore, clinical benefit could be found in the RPD group in the subgroup of well & moderate histological differentiation, preoperative CEA level less than 5·00 ng/ml, R0 resection and postoperative adjuvant chemotherapy based on DFS analysis. A multivariate Cox proportional hazards model showed that initial CA19-9≥200 U/ml, N1 stage and poor differentiation suggested poor OS. Radical resection achieved equivalent prognosis regardless of perineural invasion except for aortic nerve plexus invasion. The RPD group exhibited significantly lower locoregional and mesenteric LN recurrence rates but a higher peritoneal seeding frequency. The RPD group also exhibited significantly less back pain intensity 6 months after surgery than the SPD group. Interpretation: This study suggests that in pancreatic head cancer patients, RPD with retroperitoneal nerve resection does not provide a significant survival benefit compared with SPD. RPD can be performed safely, and be considered a treatment option with improving prognosis for patients with a low chance of systemic metastasis in pancreatic head cancer. Trial Registration: ChiCTR-TRC-12002548. Funding Statement: Sun Yat-sen University Clinical Research 5010 Program (Grant No. 2012007), China; National Natural Science Foundation of China (Grant No. 81871945). Declaration of Interests: We declare no competing interests. Ethics Approval Statement: This study was approved and overseen by the clinical ethics committee of each participating hospital.

CENTRAL INTEGRATION OF LEFT VERSUS RIGHT BARORECEPTOR AFFERENT INPUT IN FEMALE SPONTANEOUSLY HYPERTENSIVE RATS

Objective: Neuromodulation of blood pressure using baroreflex activation therapy is an effective means to control hypertension in resistant hypertensive patients. In a genetic rat model of essential hypertension, the spontaneously hypertensive rats (SHRs); we were able to demonstrate in male rats that preferential central integration of left baroreceptor afferent input exists relative to that of the right afferents. It remains unknown whether laterality also influences baroreflex responses in female SHRs. Accordingly, the differences in cardiovascular responses triggered by stimulation of the left and right baroreceptor afferents, as assessed by direct electrical stimulation of the aortic depressor nerve (ADN), were studied in female SHRs. Design and method: Pentobarbital-anesthetized female SHRs (25–29 weeks, n = 6–8) were instrumented for left and right ADN stimulation (1–40 Hz, 0.2 ms, 0.4 mA for 20 s) and recording of mean arterial pressure (MAP), heart rate (HR) and mesenteric (MVR) and femoral (FVR) vascular resistance. Female rats were also matched for the diestrus phase of the estrus cycle. Results: Both left and right ADN stimulation resulted in frequency-dependent reductions (P < 0.001) in MAP, HR, MVR and FVR. Reflex reductions in MAP, HR, MVR and FVR were comparable for both left-sided and right-sided stimulation. Conclusions: Unlike males, female SHRs express no differential central modulation of baroreceptor afferent input as evidenced by the comparable left versus right baroreflex-mediated depressor responses. Similar depressor responses in the left versus right stimulation may have been contributed to by the lack of differences in baroreflex-driven changes in HR and vascular resistance. Clinically, targeting either the left or right aortic nerves may equally provide adequate reductions in blood pressure in female hypertensive subjects. Future studies should elucidate the mechanisms responsible for sex differences in the central processing of baroreflex between male and female SHRs.

Piezo2 channel in nodose ganglia neurons is essential in controlling hypertension in a pathway regulated directly by Nedd4-2

Baroreflex plays a crucial role in regulation of arterial blood pressure (BP). Recently, Piezo1 and Piezo2, the mechanically-activated (MA) ion channels, have been identified as baroreceptors. However, the underlying molecular mechanism for regulating these baroreceptors in hypertension remains unknown. In this study, we used spontaneously hypertensive rats (SHR) and NG-Nitro-l-Arginine (L-NNA)- and Angiotensin II (Ang II)-induced hypertensive model rats to determine the role and mechanism of Piezo1 and Piezo2 in hypertension. We found that Piezo2 was dominantly expressed in baroreceptor nodose ganglia (NG) neurons and aortic nerve endings in Wistar-Kyoto (WKY) rats. The expression of Piezo2 not Piezo1 was significantly downregulated in these regions in SHR and hypertensive model rats. Electrophysiological results showed that the rapidly adapting mechanically-activated (RA-MA) currents and the responsive neuron numbers were significantly reduced in baroreceptor NG neurons in SHR. In WKY rats, the arterial BP was elevated by knocking down the expression of Piezo2 or inhibiting MA channel activity by GsMTx4 in NG. Knockdown of Piezo2 in NG also attenuated the baroreflex and increased serum norepinephrine (NE) concentration in WKY rats. Co-immunoprecipitation experiment suggested that Piezo2 interacted with Neural precursor cell-expressed developmentally downregulated gene 4 type 2 (Nedd4-2, also known as Nedd4L); Electrophysiological results showed that Nedd4-2 inhibited Piezo2 MA currents in co-expressed HEK293T cells. Additionally, Nedd4-2 was upregulated in NG baroreceptor neurons in SHR. Collectively, our results demonstrate that Piezo2 not Piezo1 may act as baroreceptor to regulate arterial BP in rats. Nedd4-2 induced downregulation of Piezo2 in baroreceptor NG neurons leads to hypertension in rats. Our findings provide a novel insight into the molecular mechanism for the regulation of baroreceptor Piezo2 and its critical role in the pathogenesis of hypertension.

Both Dietary Salt Restriction and Excess Dietary Salt Intake Exaggerate Cardiovascular Reflex Responses to Activation of the Sciatic Afferents, Aortic Depressor Nerve, and Vagal Afferents

Excess dietary salt intake exaggerates sympathetic nerve activity (SNA) and arterial blood pressure (ABP) responses to several cardiovascular reflexes in salt‐resistant animals. Recently, observational cohort studies suggest that salt restriction increases cardiovascular morbidity/mortality in several cardiovascular diseases, and the relationship between cardiovascular morbidity and dietary salt intake resembles a J‐shaped curve. However, little evidence exists to document the impact of salt restriction on cardiovascular reflex responses and whether this relationship resembles a J‐shaped curve. Thus, the aim of the present study was to determine whether salt restriction affects SNA and ABP responses to activation of sciatic afferents, the aortic depressor nerve, and cervical vagal afferents. To address this hypothesis, male Sprague‐Dawley rats were fed low (0.01% NaCl, D17010, Research diets), normal (0.1% NaCl, D17020, Research Diets), and high (4.0% NaCl, D17013, Research Diets) salt diet for 14–21 days. Then, reflex responses were measured in Inactin‐anesthetized animals. Baseline mean ABP was not different across groups (low: 112±4, normal: 101±5, high: 109±3mmHg; P>0.1 overall ANOVA). Electrical stimulation of sciatic afferents (1ms pulse, 500uA, 5s duration, 2–20Hz) produced frequency‐dependent increases in ABP. Interestingly, the pressor response to sciatic nerve stimulation was significantly greater in rats fed low and high versus normal NaCl (5Hz; low: 22±4, normal: 12±2, high: 27±2 mmHg, n=5–10, P<0.05). Activation of the aortic depressor nerve (2ms pulse, 500uA, 15s duration, 2–20Hz) produced frequency‐dependent decreases in ABP. The depressor response to aortic nerve stimulation was significantly greater in rats fed low and high versus normal NaCl (5Hz; low: −29±6, normal: −11±3, high: −29±4 mmHg; n=3–7, P<0.05). Finally, electrical stimulation of the cervical vagal afferents (1ms pulse, 500uA, 5s duration, 2–20Hz) produced frequency‐dependent decreases in ABP. Again, the depressor response to vagal nerve stimulation was significantly greater in rats fed low and high versus normal NaCl (5Hz; low: −25±3, normal: −15±2, high: −29±2 mmHg; n=5–10, P<0.05). Simultaneous recordings of lumbar, renal, and splanchnic SNA followed the changes in ABP across low, normal, and high NaCl diets. The present findings showed that both dietary salt restriction and excess dietary salt intake exaggerates sympathetic and cardiovascular responses. These cardiovascular responses to activation of sciatic afferents, the aortic depressor nerve, and vagal afferents as a function of dietary salt resemble a J‐shaped curve.Support or Funding InformationNIH R01 HL128388 (WBF, SDS)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Baroreflex Control of Heart Rate in Mice Overexpressing Human SOD1: Functional Changes in Central and Vagal Efferent Components

Excessive reactive oxygen species (ROS) (such as the superoxide radical) are commonly associated with cardiac autonomic dysfunctions. Though superoxide dismutase 1 (SOD1) overexpression may protect against ROS damage to the autonomic nervous system, superoxide radical reduction may change normal physiological functions. Previously, we demonstrated that human SOD1 (hSOD1) overexpression does not change baroreflex bradycardia and tachycardia but rather increases aortic depressor nerve activity in response to arterial pressure changes in C57B6SJL-Tg (SOD1)2 Gur/J mice. Since the baroreflex arc includes afferent, central, and efferent components, the objective of this study was to determine whether hSOD1 overexpression alters the central and vagal efferent mediation of heart rate (HR) responses. Our data indicate that SOD1 overexpression decreased the HR responses to vagal efferent nerve stimulation but did not change the HR responses to aortic depressor nerve (ADN) stimulation. Along with the previous study, we suggest that SOD1 overexpression preserves normal baroreflex function but may differentially alter the functions of the ADN, vagal efferents, and central components. While SOD1 overexpression likely enhanced ADN function and the central mediation of bradycardia, it decreased vagal efferent control of HR.

SOD1 Overexpression Preserves Baroreflex Control of Heart Rate with an Increase of Aortic Depressor Nerve Function

Overproduction of reactive oxygen species (ROS), such as the superoxide radical (O2 ∙−), is associated with diseases which compromise cardiac autonomic function. Overexpression of SOD1 may offer protection against ROS damage to the cardiac autonomic nervous system, but reductions of O2 ∙− may interfere with normal cellular functions. We have selected the C57B6SJL-Tg (SOD1)2 Gur/J mouse as a model to determine whether SOD1 overexpression alters cardiac autonomic function, as measured by baroreflex sensitivity (BRS) and aortic depressor nerve (ADN) recordings, as well as evaluation of baseline heart rate (HR) and mean arterial pressure (MAP). Under isoflurane anesthesia, C57 wild-type and SOD1 mice were catheterized with an arterial pressure transducer and measurements of HR and MAP were taken. After establishing a baseline, hypotension and hypertension were induced by injection of sodium nitroprusside (SNP) and phenylephrine (PE), respectively, and ΔHR versus ΔMAP were recorded as a measure of baroreflex sensitivity (BRS). SNP and PE treatment were administered sequentially after a recovery period to measure arterial baroreceptor activation by recording aortic depressor nerve activity. Our findings show that overexpression of SOD1 in C57B6SJL-Tg (SOD1)2 Gur/J mouse preserved the normal HR, MAP, and BRS but enhanced aortic depressor nerve function.

Differential contribution of afferent and central pathways to the development of baroreflex dysfunction in chronic kidney disease

The effects of chronic kidney disease on baroreflex control of renal sympathetic nerve activity (RSNA) and deficits in afferent and central components of the baroreflex were studied in juvenile and adult male Lewis Polycystic Kidney (LPK) and control Lewis rats under anesthesia (n=35). Blood pressure (BP), heart rate (HR), aortic depressor nerve activity (ADNA), and RSNA were determined after pharmacological manipulation of BP. Responses to ADN stimulation (4.0 V, 2.0 ms, 1-24 Hz) were determined, and the aortic arch was collected for histomorphometry. In juvenile LPK versus age-matched Lewis rats, gain of RSNA (-1.5±0.2 versus -2.8±0.2%/mm Hg; P<0.05) and ADNA (2.5±0.3 versus 5.0±0.6%/mm Hg; P<0.05), but not HR barocurves, were reduced. BP, HR, and RSNA responses to ADN stimulation were normal or enhanced in juvenile LPK. In adult LPK versus age-matched Lewis, the gain and range of RSNA (gain: -1.2±0.1 versus -2.2±0.2%/mm Hg, range: 62±8 versus 98±7%) and HR (gain: -0.7±0.1 versus -3.5±0.7 bpm/mm Hg, range: 44±8 versus 111±19 bpm) barocurves were reduced (P<0.05). The gain and range of the ADNA barocurves were also reduced in adult LPK versus Lewis [1.5±0.4 versus 5.2±1.1 (%/mm Hg) and 133±35 versus 365±61 (%) P<0.05] and correlated with aortic arch vascular remodeling. BP, HR, and RSNA responses to ADN stimulation were significantly reduced in adult LPK. Our data demonstrate a deficit in the afferent component of the baroreflex that precedes the development of impaired central regulation of RSNA and HR in chronic kidney disease, and that progressive impairment of both components is associated with marked dysfunction of the baroreflex pathway.

Open-loop characteristics of the arterial baroreflex after blockade of unmyelinated baroreceptors with resiniferatoxin.

Arterial baroreceptors can be divided into two categories dependent on whether their axons are myelinated (A-fiber) or unmyelinated (C-fiber). We investigated the effect of periaxonal resiniferatoxin (RTX), a blocker of C-fiber baroreceptor activity, on the open-loop static characteristics of the arterial baroreflex. The baroreceptor region of the right aortic depressor nerve was isolated, and intra-baroreceptor region pressure (BRP) was changed from 60 to 180 mm Hg in 10 anesthetized Sprague-Dawley rats. Open-loop static characteristics of the neural arc from BRP to efferent sympathetic nerve activity (SNA), peripheral arc from SNA to arterial pressure (AP), and total reflex arc from BRP to AP were estimated. Although blocking C-fiber activity with RTX resulted in a lower response range (33.7±4.6% and 49.4±4.8%, P<0.01) and higher minimum SNA (78.0±4.7% and 53.6±5.0%, P<0.001) of the steady-state neural arc, the peak SNA response to BRP was greater at a BRP of 160 mm Hg (-37.87±5.83% and -26.28±4.90%, P=0.01). RTX also resulted in a lower response range (27.8±5.0 mm Hg and 40.9±5.2 mm Hg, P<0.01) and higher minimum AP (92.4±4.7 mm Hg and 79.1±4.9 mm Hg, P<0.001) of the total reflex arc. Despite these changes, the maximum slope of the neural arc and the maximum gain of the total reflex arc did not differ significantly after RTX. These data suggest that A-fiber baroreceptors can regulate AP and maintain the maximum gain when systemic AP is around the normal operating range. In contrast, C-fiber baroreceptors are critically important for reductions in SNA and AP when systemic AP is raised above the normal operating range.

Abstract 458: The Baroreflex Modulates The Anti-inflammatory Response Caused By Intravenous Lipopolysaccharide (lps)

Activation of the baroreflex elicits parasympathetic overactivity combined with sympathetic inhibition. The effect of electrical stimulation (ES) of the aortic depressor nerve (ADN) on the hemodynamic and inflammatory responses elicited by LPS (1.5 mg/kg iv) was examined in conscious rats. Rats were divided into 4 groups: 1) SALINE (n=6); 2) ES+SALINE (n=5); 3) LPS (n=7); 4) ES+LPS (n=5). Under ketamine/xylazine anesthesia the subjects were implanted with a pair of electrodes around the ADN and catheters into the femoral artery and jugular vein. On the next day, mean arterial pressure (MAP) and heart rate (HR) were recorded. ES (0.5mA; 0.25ms; 15Hz) was performed during 60 min after saline or LPS administration. Plasma was collected 1, 2 and 3h after saline or LPS administration, while the spleen and hypothalamus were collected after 3h to quantify interleukin (IL10) and tumor necrosis factor (TNFα). ES in the SALINE group promoted a prompt (5s) fall in MAP (-16± 5 mmHg) and HR (-39±12 bpm) followed by complete recovery of the MAP (7± 3 mmHg) and partial recovery of the HR (-8 ± 10 bpm) after 1h. In the LPS group ES promoted a prompt (5s) fall in MAP (-19± 6 mmHg) and HR (-40± 15 bpm) followed by complete recovery of the MAP (1 ± 3 mmHg) combined with significant tachycardia (30± 14bpm). Plasma TNFα increased 1h after LPS (400±78 vs 11±4 pg/μL from SALINE) followed by a decrease 2h (80±14 pg/μL) and 3h (34,4±5 pg/μL). After LPS the ES augmented IL-10 in the spleen (344±37 vs. 518± 79 pg/μL from ES+LPS) and decreased in the hypothalamus (73±8 vs 57±5 pg/μL from ES+LPS); while after saline ES increased IL-10 in the spleen (63±18 vs 314±99 pg/μL from ES+SALINE) but did not change in the hypothalamus (84±10 vs 97±17 pg/μL from ES+SALINE). After LPS administration ES did not change TNFα in the spleen (428±50 vs 450±98 pg/μL from ES+LPS) and hypothalamus (48±4 vs 38±5 pg/μL from ES+LPS); likewise in the spleen (285±38 vs 286±83 pg/μL from ES+SALINE) and hypothalamus (51,7±6 vs 54±8 pg/μL from ES+SALINE) after saline administration. These findings indicate that electrical activation of the baroreflex in conscious rats was capable to modulate the inflammatory response, promoting an increase of anti-inflammatory mediators in the LPS model of inflammation.

Discharges of aortic and carotid sinus baroreceptors during spontaneous motor activity and pharmacologically evoked pressor interventions.

Our laboratory has demonstrated that the cardiomotor component of aortic baroreflex is temporarily inhibited at the onset of spontaneous motor activity in decerebrate cats, without altering carotid sinus baroreflex. A reason for this dissociation may be attributed to a difference in the responses between aortic nerve activity (AoNA) and carotid sinus nerve activity (CsNA) during spontaneous motor activity. The stimulus-response curves of AoNA and CsNA against mean arterial blood pressure (MAP) were compared between the pressor interventions evoked by spontaneous motor activity and by intravenous administration of phenylephrine or norepinephrine, in which the responses in heart rate (HR) were opposite (i.e., tachycardia vs. baroreflex bradycardia), despite the identical increase in MAP of 34-40 mmHg. In parallel to the pressor response, mean AoNA and CsNA increased similarly by 78-81 and by 88 % of the baseline control, respectively, irrespective of whether the pressor response was evoked by spontaneous motor activity or by a pharmacological intervention. The slope of the stimulus-response curve of the mean AoNA became greater (P < 0.05) during spontaneous motor activity as compared to the pharmacological intervention. On the other hand, the stimulus-response curve of the mean CsNA and its slope were equal (P > 0.05) between the two pressor interventions. Furthermore, the slopes of the stimulus-response curves of both diastolic AoNA and CsNA (defined as the minimal value within a beat) exhibited a greater increase during spontaneous motor activity. All differences in the slopes of the stimulus-response curves were abolished by restraining HR at the intrinsic cardiac frequency. In conclusion, mean mass activities of both aortic and carotid sinus baroreceptors are able to encode the beat-by-beat changes in MAP not only at rest but also during spontaneous motor activity and spontaneous motor activity-related reduction of aortic baroreceptor activity is denied accordingly.

Hemodynamic responses to electrical stimulation of carotid sinus in conscious rats (1169.15)

Carotid sinus (CS) is an important site of arterial baroreceptors and its electrical stimulation has been used to treat patients with resistant hypertension. This study evaluated hemodynamic responses to electrical stimulation of CS in normal conscious rats. Rats (N=5) had left carotid sinus (CS) carefully isolated, equipped with a pair of electrodes and insulated with a layer of Kwik sil. A catheter was introduced into abdominal aorta throughout laparotomy and exteriorized in the back of the neck, together with the CS electrodes. After two days, conscious rats, under continuous recording of AP and HR, had their CS electrically stimulated, during 20 s, with square pulses of 2 ms, 1 mA and 15, 30, 60 and 90 Hz. Rats showed a basal arterial pressure (AP) and heart rate (HR) of 108±3 mmHg and 353±9 bpm. The recorded voltage of the 1 mA pulses allowed the calculation of carotid sinus resistance that ranged from 4.3 to 7.2 kΩ. A fall in mean AP (Δ=19±2, 28±7, 31±4 and 39±4 mmHg) was observed 5‐9 s after the onset of carotid sinus stimulation with 15, 30, 60 and 90 Hz, respectively. A slight and non‐statistically significant fall in HR (9‐35 bpm) was also observed immediately (1‐3 s) after the onset of CS stimulation. In contrast to electrical stimulation of aortic nerve in conscious rats, currently performed in our lab, CS stimulation affected mainly vascular resistance, once only a slight and non‐significant effect was seem in HR.Grant Funding Source: Supported by CAPES, FAPESP, CNPq.

Signal transduction of aortic and carotid sinus baroreceptors is not modified by central command during spontaneous motor activity in decerebrate cats

Our laboratory has suggested that central command provides selective inhibition of the cardiomotor component of aortic baroreflex at the start of exercise, preserving carotid sinus baroreflex. It is postulated that central command may modify the signal transduction of aortic baroreceptors, so as to decrease aortic baroreceptor input to the cardiovascular centers, and, thereby, can cause the selective inhibition of aortic baroreflex. To test the hypothesis, we directly analyzed the responses in multifiber aortic nerve activity (AoNA) and carotid sinus nerve activity (CsNA) during spontaneous motor activity in decerebrate, paralyzed cats. The increases of 62-104% in mean AoNA and CsNA were found during spontaneous motor activity, in proportion to a rise of 35 ± 3 mmHg (means ± SE) in mean arterial blood pressure (MAP), and had an attenuating tendency by restraining heart rate (HR) at the lower intrinsic frequency of 154 ± 6 beats/min. Brief occlusion of the abdominal aorta was conducted before and during spontaneous motor activity to produce a mechanically evoked increase in MAP and, thereby, to examine the stimulus-response relationship of arterial baroreceptors. Although the sensitivity of the MAP-HR baroreflex curve was markedly blunted during spontaneous motor activity, the stimulus-response relationships of AoNA and CsNA were not influenced by spontaneous motor activity, irrespective of the absence or presence of the HR restraint. Thus, it is concluded that aortic and carotid sinus baroreceptors can code beat-by-beat blood pressure during spontaneous motor activity in decerebrate cats and that central command is unlikely to modulate the signal transduction of arterial baroreceptors.

Acute Experimental Diabetes and Aortic Depressor Nerve Ultrastructural Morphometry: Effects of Insulin Treatment

Previous results from our laboratory showed myelinated fiber alterations in the aortic depressor nerve (ADN) from rats rendered diabetic for 15 days (short term). The ultrastructure and the acute effect of insulin treatment, particularly in the unmyelinated fibres of this nerve, were not yet investigated in this experimental model, being the aim of this work. Wistar rats received a single intravenous injection of streptozotocin (STZ – 40 mg/Kg) 15 days (N=7) before the experiments. Control animals (N=7) received vehicle (citrate buffer) and treated animals (n=6) received a subcutaneous injection of insulin (10 IU/Kg) on a daily basis beginning 3 days after the STZ injection. Under pentobarbital anaesthesia the ADNs were isolated and had their spontaneous activity recorded. Afterwards, proximal and distal segments of the nerves were prepared for transmission electron microscopy studies. Morphometry of the myelinated and unmyelinated fibres was carried out with the aid of computer software. Nerves from diabetic animals showed some unmyelinated fiber enlargement with signs of swelling and degeneration, and the Schwann cells mitochondria presented enlargement and clustering formation. On treated animals, an important thickening of the Schwann cells’ basement membrane was present and some myelinated fibers showed thin myelin sheaths. The endoneural blood vessels ultrastrucutre was preserved in all nerves from the three experimental groups. Myelinated axon area and diameter was significantly larger on insulin treated animals, compared to both, controls and acute diabetic groups, on both segments. No other differences were observed between groups for the myelinated or unmyelinated fibers morphometric parameters. Investigation of experimental models of diabetes in early or acute stage of the disease provide relevant information on the physiopathological mechanisms involved in the alterations present in patients with chronic diabetes and its complications. We suggest that insulin treatment alone might have a role on the morphological alterations present in diabetic neuropathy.

Orexin-neuromodulated cerebellar circuit controls redistribution of arterial blood flows for defense behavior in rabbits

We investigated a unique microzone of the cerebellum located in folium-p (fp) of rabbit flocculus. In fp, Purkinje cells were potently excited by stimulation of the hypothalamus or mesencephalic periaqueductal gray, which induced defense reactions. Using multiple neuroscience techniques, we determined that this excitation was mediated via beaded axons of orexinergic hypothalamic neurons passing collaterals through the mesencephalic periaqueductal gray. Axonal tracing studies using DiI and biotinylated dextran amine evidenced the projection of fp Purkinje cells to the ventrolateral corner of the ipsilateral parabrachial nucleus (PBN). Because, in defense reactions, arterial blood flow has been known to redistribute from visceral organs to active muscles, we hypothesized that, via PBN, fp adaptively controls arterial blood flow redistribution under orexin-mediated neuromodulation that could occur in defense behavior. This hypothesis was supported by our finding that climbing fiber signals to fp Purkinje cells were elicited by stimulation of the aortic nerve, a high arterial blood pressure, or a high potassium concentration in muscles, all implying errors in the control of arterial blood flow. We further examined the arterial blood flow redistribution elicited by electric foot shock stimuli in awake, behaving rabbits. We found that systemic administration of an orexin antagonist attenuated the redistribution and that lesioning of fp caused an imbalance in the redistribution between active muscles and visceral organs. Lesioning of fp also diminished foot shock-induced increases in the mean arterial blood pressure. These results collectively support the hypothesis that the fp microcomplex adaptively controls defense reactions under orexin-mediated neuromodulation.

Plasma leptin inhibits the response of nucleus of the solitary tract neurons to aortic baroreceptor stimulation

Leptin receptors have been identified within the nucleus of the solitary tract (NTS) and leptin injections into the caudal NTS inhibit the baroreceptor reflex. However, whether plasma leptin alters the discharge of NTS neurons mediating aortic baroreceptor reflex activity is not known. A series of electrophysiological single unit recording experiments was done in the urethane-chloralose anesthetized, paralyzed and artificially ventilated Wistar and Zucker obese rat with either their neuroaxis intact or with mid-collicular transections. Single units in NTS antidromically activated by electrical stimulation of depressor sites in the caudal ventrolateral medulla (CVLM) were found to display a cardiac cycle-related rhythmicity. These units were tested for their responses to stimulation of the aortic depressor nerve (ADN) and intra-carotid injections of leptin (50-200ng/0.1ml). Of 63 single units tested in NTS, 33 were antidromically activated by stimulation of CVLM depressor sites and 18 of these single units responded with a decrease in discharge rate after intracarotid injections of leptin. Thirteen of these leptin responsive neurons (∼72%) were excited by ADN stimulation. Furthermore, the excitatory response of these single units to ADN stimulation was attenuated by about 50% after the intracarotid leptin injection. Intracarotid injections of leptin (200ng/0.1ml) in the Zucker obese rat did not alter the discharge rate of NTS-CVLM projecting neurons. These data suggest that leptin exerts a modulatory effect on brainstem neuronal circuits that control cardiovascular responses elicited during the reflex activation of arterial baroreceptors.

Leptin in nucleus of the solitary tract alters the cardiovascular responses to aortic baroreceptor activation

Recent data suggests that neurons expressing the long form of the leptin receptor form at least two distinct groups within the caudal nucleus of the solitary tract (NTS): a group within the lateral NTS (Slt) and one within the medial (Sm) and gelantinosa (Sg) NTS. Discrete injections of leptin into Sm and Sg, a region that receives chemoreceptor input, elicit increases in arterial pressure (AP) and renal sympathetic nerve activity (RSNA). However, the effect of microinjections of leptin into Slt, a region that receives baroreceptor input is unknown. Experiments were done in the urethane-chloralose anesthetized, paralyzed and artificially ventilated Wistar or Zucker obese rat to determine leptin's effect in Slt on heart rate (HR), AP and RSNA during electrical stimulation of the aortic depressor nerve (ADN). Depressor sites within Slt were first identified by the microinjection of l-glutamate (Glu; 0.25M; 10nl) followed by leptin microinjections. In the Wistar rat leptin microinjection (50ng; 20nl) into depressor sites within the lateral Slt elicited increases in HR and RSNA, but no changes in AP. Additionally, leptin injections into Slt prior to Glu injections at the same site or to stimulation of the ADN were found to attenuate the decreases in HR, AP and RSNA to both the Glu injection and ADN stimulation. In Zucker obese rats, leptin injections into NTS depressor sites did not elicit cardiovascular responses, nor altered the cardiovascular responses elicited by stimulation of ADN. Those data suggest that leptin acts at the level of NTS to alter the activity of neurons that mediate the cardiovascular responses to activation of the aortic baroreceptor reflex.

Ultrastructural Morphometry of the Aortic Depressor Nerves and Extrinsic Renal Nerves: Similarities and Differences between Mice and Rats

Anatomical and physiological aspects involving rats and mice have shown similarities and differences between these experimental animal models. In cardiovascular physiology research, rats are being substituted by mice since mice are more susceptible to genetic manipulation. Nevertheless, little is known about mice normal anatomy and/or physiology to allow the correct interpretation of altered responses on genetically manipulated animals. We compared morphometric ultrastructural parameters of the aortic depressor nerve (ADN) and extrinsic renal nerve (SRN) between Wistar rats and C57BL/6J mice. After spontaneous activity recordings together with the arterial pressure pulse, ADN and SRN were dissected and prepared for light and transmission electron microscopy. Morphometry was performed with an image analysis software and took into consideration the fascicle area and diameter, myelinated and unmyelinated fiber number, density, area and diameter, myelin sheath area and the g-ratio. Comparisons were made for the same nerve between mice and rats and differences were considered significant when p<0.05. Both nerves were large in rats compared to mice, as were the myelinated fibers. On the other hand, unmyelinated fibers density, size and distributions were not different between species. These results suggest physiological differences on the fast conduction fibers between species, with possible different functional role. Morphological comparisons of the quantitative composition of peripheral nerves in different species and strains are very rare. Our study contributes to a morphological understanding of important nerves related to cardiovascular reflexes in two animal models. It also presents, for the first time, ultrastructural morphometric characteristics of the ADN in mice. This data provide a morphological basis for further studies involving functional investigations for reflex regulation of circulation in experimental model of hypertension and to improved cardiovascular system knowledge.