Bilateral Cervical Vagotomy Research Articles

Co-dependence of the neural and humoral pathways in the mechanism of remote ischemic conditioning.

The cardioprotection afforded by remote ischaemic conditioning (RIC) is mediated via a complex mechanism involving sensory afferent nerves, the vagus nerve, and release of a humoral blood-borne factor. However, it is unknown whether release of the protective factor depends on vagal activation or occurs independently. This study aimed to evaluate the co-dependence of the neural and humoral pathways of RIC, focussing on the vagus nerve and intrinsic cardiac ganglia. In the first study, anesthetised rats received bilateral cervical vagotomy or sham-surgery immediately prior to RIC (4 × 5 min limb ischemia–reperfusion) or sham-RIC. Venous blood plasma was dialysed across a 12–14 kDa membrane and dialysate perfused through a naïve-isolated rat heart prior to 35-min left anterior descending ischemia and 60-min reperfusion. In the second study, anesthetised rats received RIC (4 × 5-min limb ischemia–reperfusion) or control (sham-RIC). Dialysate was prepared and perfused through a naïve-isolated rat heart in the presence of the ganglionic blocker hexamethonium or muscarinic antagonist atropine, prior to ischemia–reperfusion as above. Dialysate collected from RIC-treated rats reduced infarct size in naïve rat hearts from 40.7 ± 6.3 to 23.7 ± 3.1 %, p < 0.05. Following bilateral cervical vagotomy, the protection of RIC dialysate was abrogated (42.2 ± 3.2 %, p < 0.05 vs RIC dialysate). In the second study, the administration of 50-μM hexamethonium (45.8 ± 2.5 %) or 100-nM atropine (36.5 ± 3.4 %) abrogated the dialysate-mediated protection. Release of a protective factor following RIC is dependent on prior activation of the vagus nerve. In addition, this factor appears to induce cardioprotection via recruitment of intrinsic cardiac ganglia.Electronic supplementary materialThe online version of this article (doi:10.1007/s00395-016-0568-z) contains supplementary material, which is available to authorized users.

Vagus nerve electrical stimulation inhibits serum levels of S100A8 protein in septic shock rats.

The vagus nerve and the released acetylcholine exert anti-inflammatory effects and inhibit septic shock. However, their detailed mechanisms remain to be elucidated. The present study aimed to investigate the effects of vagus nerve electrical stimulation on serum S100A8levels in septic shock rats. A total of36male Sprague-Dawley rats were randomly divided into six equal groups: i)Sham group, receiving sham operation; ii)CLP group, subjected to cecal ligation and puncture (CLP) to establish a model of polymicrobial sepsis; iii)VGX group, subjected to CLP and bilateral cervical vagotomy; iv)STM group, subjected to CLP, bilateral cervical vagotomy and electrical stimulation on the left vagus nerve trunk; v)α‑bungarotoxin (BGT) group was administered α‑BGT prior to electrical stimulation; vi)Anti‑receptor for advanced glycation end products (RAGE) group, administered intraperitoneal injection of anti‑RAGE antibody prior to electrical stimulation. The right carotid artery was cannulated to monitor mean artery pressure (MAP). The serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels were measured to assess the liver function. Serum S100A8and advanced glycation end product (AGE) levels were measured using enzyme‑linked immunosorbent assays. The expression of hepatic RAGE was determined by western blotting. The present study revealed that Sprague‑Dawley rats exhibited progressive hypotension and significantly increased serum AST and ALT levels following CLP challenge compared with the sham group. The levels of S100A8and AGEs, and the protein expression of hepatic RAGE were significantly increased following CLP compared with the sham group. Vagus nerve electrical stimulation significantly prevented the development of CLP‑induced hypotension, alleviated the hepatic damage, reduced serum S100A8 and AGEs production, and reduced the expression of hepatic RAGE. The inhibitory effect of vagus nerve electrical stimulation was reversed by pre-treatment with α-BGT and enhanced by pre-treatment with anti-RAGE antibody. In conclusion, vagus nerve electrical stimulation may produce a protective effect on septic shock in rats by decreasing the production of S100A8.

Cervical vagotomy increased the distal colon distention to urinary bladder inhibitory reflex in male rats.

Many studies have demonstrated the convergence of vagal inputs into brainstem centers with inputs from the urinary bladder and colon, as well as the convergence of vagal inputs into other centers controlling the urinary bladder and colon reflexes. However, the effect of the vagal inputs on the interaction between the urinary bladder and other pelvic organs has not been studied. In this study, the effect of bilateral cervical vagotomy on the distal colon to urinary bladder reflex was examined. Changes to cystometry parameters in response to increased distal colon distensions (1, 2, and 3 ml) were tested in urethane-anesthetized male rats with or without bilateral cervical vagotomy. In animals with intact vagus nerves, 1 and 2 ml distal colon distentions had no significant effects on micturition frequency; however, 3 ml distal colon distention significantly decreased the frequency of micturition cycles. Also, 3 ml distal colon distention inhibited micturition cycles in 37.5 % of these animals. On the other hand, following cervical vagotomy, 1 ml distal colon distention was enough to significantly decrease the frequency of micturition cycles and to inhibit the cycles in 75 % of the animals. These results demonstrate the presence of supraspinal inhibitory regulation, via the vagus nerve, over the distal colon to urinary bladder inhibitory reflex.

NDP-α-MSH attenuates heart and liver responses to myocardial reperfusion via the vagus nerve and JAK/ERK/STAT signaling

Melanocortin peptides afford cardioprotection during myocardial ischemia/reperfusion via janus kinases (JAK), extracellular signal-regulated kinases (ERK) and signal transducers/activators of transcription (STAT) pathways. Here we investigated whether melanocortin-induced modulation of the JAK/ERK/STAT signaling occurs via the cholinergic anti-inflammatory pathway, focusing our study on cardiac and hepatic responses to prolonged myocardial ischemia/reperfusion. Ischemia was produced in rats by ligature of the left anterior descending coronary artery for 30min; effects of ischemia/reperfusion were evaluated using Western blot of heart and liver proteins. Intravenous treatment, during coronary artery occlusion, with the melanocortin analog (Nle4, D-Phe7)α-melanocyte-stimulating hormone (NDP-α-MSH) induced a left ventricle up-regulation of the cardioprotective transcription factors pJAK2, pERK1/2 and pTyr-STAT3 (JAK-dependent), and a reduction in the levels of the inflammatory mediators tumor necrosis factor-α (TNF-α) and pJNK (a transcription factor also involved in apoptosis), as assessed at the end of the 2-h reperfusion period. Further, these beneficial effects of NDP-α-MSH were associated with heart over-expression of the pro-survival proteins heme oxygenase-1 (HO-1) and Bcl-XL, and decrease of ventricular arrhythmias and infarct size. In the liver NDP-α-MSH induced a decrease in the pJAK2 and pTyr-STAT3 levels, and strongly reduced pERK1/2 expression. In the liver of ischemic rats NDP-α-MSH also blunted pJNK activity and TNF-α expression, and up-regulated Bcl-XL. Bilateral cervical vagotomy prevented all effects of NDP-α-MSH, both in the heart and liver. These results indicate that melanocortins inhibit heart and liver damage triggered by prolonged myocardial ischemia/reperfusion likely, as main mechanism, via the vagus nerve-mediated modulation of the JAK/STAT/ERK signaling pathways.

Effect of bilateral cervical vagotomy on microglial activation in spinal cord in a rat model of persistent postoperative pain

Objective To investigate the effect of bilateral cervical vagotomy on microglial activation in spinal cord in a rat model of persistent postoperative pain evoked by skin/muscle incision and retraction (SMIR). Methods Thirty six male Sprague-Dawley rats were randomly divided into three groups (n= 12 each group): group sham operation, group SMIR, and group SMIR+ bilateral cervical vagotomy (SV). The rat model of persistent postoperative pain evoked by SMIR was established according to the method described by Flatter. Pain behavior was assessed by paw mechanical withdrawal threshold (MWT) to von Frey filament stimulation at 1, 3, and 5 days after operation. Four animals were sacrificed at each time point in each group to detect the expression of Iba-1 (a specific marker of microglia) in the spinal dorsal horn with immunofluorescence and the microglia was counted. Results MWT was significantly decreased at T1-5 in SMIR and SV (10.3±0.6, 9.7±0.8, 9.6±0.5; 8.0±0.7, 7.7±0.4, 7.6±0.3), while the expression of Iba-1 and microglia counts in the spinal dorsal horn were significantly increased at T1-5 in SMIR and SV (1428±134, 1245±129, and 1001±117; 8.0±0.7, 7.7±0.4, and 7.6±0.3; 187±13, 164±11, and 142±14; and 241±21, 230±21, and 202±19). In group SV as compared to group SMIR, MWT was significantly decreased at T1-5, while the expression of Iba-1 and microglia counts in the spinal dorsal horn were significantly increased at T1-5. Conclusions Vagus nerve plays an important role in microglial activation in spinal cord in a rat model of persistent postoperative pain evoked by skin/muscle incision and retraction. Key words: Vagotomy; Pain, postoperative; Microglia; Rats, Sprague-Dawley

Evidence for the gastric cytoprotective effect of centrally injected agmatine.

Agmatine (decarboxylated arginine) exerts cytoprotective action in several tissues, such as in the brain, heart or kidneys, but there is still controversy over the effects of agmatine on the gastric mucosa. The aim of the present study was to reveal the potential gastroprotective action of agmatine by using an acid-independent ulcer model to clarify which receptors and peripheral factors are involved in it. Gastric mucosal damage was induced by acidified ethanol. Mucosal levels of calcitonin gene-related peptide (CGRP) and somatostatin were determined by radioimmunoassay. For analysis of gastric motor activity the rubber balloon method was used. It was found that agmatine given intracerebroventricularly (i.c.v., 0.044-220 nmol/rat) significantly inhibited the development of ethanol-induced mucosal damage, while in the case of intraperitoneal injection (0.001-50mg/kg i.p.) it had only a minor effect. The central gastroprotective action of agmatine was completely antagonized by mixed alpha2-adrenoceptor and imidazoline I1 receptor antagonists (idazoxan, efaroxan), but only partially by yohimbine (selective alpha2-adrenoceptor antagonist) and AGN 192403 (selective I1 receptor ligand, putative antagonist). It was also inhibited by the non-selective opioid-receptor antagonist naloxone and the selective δ-opioid receptor antagonist naltrindole, but not by β-funaltrexamine and nor-Binaltorphimine (selective μ- and κ-opioid receptor antagonists, respectively). Furthermore, the effect of agmatine was antagonized by bilateral cervical vagotomy and by pretreatment with indomethacin and NG-nitro-l-arginine. Agmatine also reversed the ethanol-induced reduction of gastric mucosal CGRP and somatostatin content, but did not have any significant effect on gastric motor activity. These results indicate that agmatine given centrally induces gastric cytoprotection, which is mediated by central imidazoline I1 receptors, alpha2-adrenoceptors and δ-opioid receptors. Activation of these receptors induces the release of different mucosal protective factors, such as NO, prostaglandins, CGRP and somatostatin by a vagal-dependent mechanism. Alterations of gastric motility are not likely to contribute to the observed protective effect.

Electroacupuncturing at Zusanli point (ST36) attenuates pro-inflammatory cytokine release and organ dysfunction by activating cholinergic anti-inflammatory pathway in rat with endotoxin challenge.

It is considered that the pathogenesis is closely related to an excessive production of pro-inflammatory cytokines caused by bacterial toxins and an imbalance between pro-inflammatory and anti-inflammatory mediators. This work investigates the effect of electro-acupuncturing (EA), at Zusanli point (ST36) on plasma cytokine release and organ dysfunction and their mechanism in conscious rats with endotoxin challenge. EA at Zusanli points obviously lowered the elevated levels of plasma TNF-α, and attenuated changes in parameters relevant to various organ functions at 2 h after LPS challenge. α-BGT injection or bilateral cervical vagotomy could weaken or eliminate the effects of EA, and further aggravated the elevated levels of pro-inflammatory cytokines and organ dysfunction. The results suggested that EA at Zusanli points significantly reduced the release of pro-inflammatory cytokines and organ dysfunction after LPS challenge by activating cholinergic anti-inflammatory pathway.

Activated pathway from the parabrachial nucleus to rostal ventrolateral medulla during stimulation of cardiac sympathetic afferents (686.8)

Our previous studies have shown that stimulation of cardiac sympathetic afferents activates pontine nuclei like the parabrachial nucleus (PBN) and locus coeruleus (LC) as well as rostral ventrolateral medulla (rVLM), a critical area that controls cardiac sympathoexcitatory responses. However, there is no information on direct influence of pontine nuclei on the rVLM during stimulation of cardiac sympathetic afferents. Thus, the present study evaluated neuronal activation of the pontine nuclei in response to stimulation of cardiac sympathetic afferents, with regard to their projections to the rVLM. Seven to ten days after unilateral microinjection of rodamine‐conjugated microspheres retrograde tracer (100 nl) into the rVLM, rats with bilateral cervical vagotomy were subjected to epicardial application of α, β‐meATP (1.1 to 6.5 mM, in 40 µl) or 0.9% normal saline for six times over 90 min. Repetitive increases in blood pressure were observed in the rats following repeated stimulation of the heart with α, β‐meATP, but not with normal saline. Perikarya containing the microsphere tracer were found in the PBN and, but not in the LC of both groups. Compared to three controls, c‐Fos immunoreactivity and neurons double‐labeled with c‐Fos, an immediate early gene and the tracer were more frequently found in the PBN, but not LC of rats with cardiac stimulation (n=5), particularly in the lateral PBN and Kölliker‐Fuse nucleus. These results suggest that PBN neurons responsive to cardiac sympathetic afferent stimulation frequently project directly to the rVLM, which is known to participate in processing of excitatory cardiovascular responses during stimulation of cardiac sympathetic afferents.Grant Funding Source: NIH grant HL‐66217

Reflex control of inflammation by sympathetic nerves, not the vagus

We investigated a neural reflex that controls the strength of inflammatory responses to immune challenge - the inflammatory reflex. In anaesthetized rats challenged with intravenous lipopolysaccharide (LPS, 60μgkg(-1)), we found strong increases in plasma levels of the key inflammatory mediator tumour necrosis factor α (TNFα) 90min later. Those levels were unaffected by previous bilateral cervical vagotomy, but were enhanced approximately 5-fold if the greater splanchnic sympathetic nerves had been cut. Sham surgery had no effect, and plasma corticosterone levels were unaffected by nerve sections, so could not explain this result. Electrophysiological recordings demonstrated that efferent neural activity in the splanchnic nerve and its splenic branch was strongly increased by LPS treatment. Splenic nerve activity was dependent on inputs from the splanchnic nerves: vagotomy had no effect on the activity in either nerve. Together, these data demonstrate that immune challenge with this dose of LPS activates a neural reflex that is powerful enough to cause an 80% suppression of the acute systemic inflammatory response. The efferent arm of this reflex is in the splanchnic sympathetic nerves, not the vagi as previously proposed. As with other physiological responses to immune challenge, the afferent pathway is presumptively humoral: the present data show that vagal afferents play no measurable part. Because inflammation sits at the gateway to immune responses, this reflex could play an important role in immune function as well as inflammatory diseases.

Angiotensin II-induced activation of central AT1 receptors exerts endocannabinoid-mediated gastroprotective effect in rats

The aim of the present study was to analyze whether angiotensin II via the endocannabinoid system can induce gastric mucosal protection, since transactivation of cannabinoid CB1 receptors by angiotensin AT1 receptor in CHO cells was described. Experimental ulcer was induced by acidified ethanol given orally in male Wistar rats, CB1(+/+) wild type and CB1(−/−) knockout mice. The compounds were administered intracerebroventricularly. It was found, that 1. Angiotensin II inhibited the ethanol-induced gastric lesions (11.9–191pmol); the effect of angiotensin II (191pmol) was inhibited by the CB1 receptor inverse agonist AM 251 (1.8nmol) and the inhibitor of diacylglycerol lipase (DAGL), tetrahydrolipstatin (0.2nmol). 2. Angiotensin II exerted gastroprotection in wild type, but not in CB1(−/−) mice. 3. The gastroprotective effect of angiotensin II (191pmol) was reduced by atropine (1mg/kg i.v.) and bilateral cervical vagotomy. In conclusion, stimulation of central angiotensin AT1 receptors via activation of cannabinoid CB1 receptors induces gastroprotection in a DAGL-dependent and vagus-mediated mechanism.

Retraction.

Early life exposure to environmentally relevant ozone (O 3 ) may have long term consequences for asthma susceptibility. Earlier studies showed that mice exposed to intermittent, repeated O 3 during early life (1 ppm x 3h/day, 3 days/week x 4 weeks) demonstrated airway hyperresponsiveness (AHR) to inhaled methacholine challenge compared with filtered air controls at adult age (4 wk), and after an additional 4 wk recovery period (age 8wk). To determine the mechanism, we exposed mice to filtered air or O 3 as described above beginning at birth. After two weeks recovery in air, we measured ex vivo tracheal smooth muscle mechanics in response to acetylcholine and electrical field stimulation. We found significant but relatively modest impairment of relaxation in ozone exposed mice. In vivo measurements of respiratory system mechanics using forced oscillometry in response to nebulized methacholine showed persistence of O 3 -exposure induced AHR that was substantially ablated in mice that had undergone: 1) pre-treatment with muscarinic receptor agonist pilocarpine, or 2) bilateral cervical vagotomy, just before undergoing forced oscillometry. Separate treatment groups that received intravenous acetylcholine in place of nebulized methacholine showed no O 3 effect on AHR. Bronchoalveolar lavage of mice after two or four weeks recovery following air or O 3 exposure showed elevations of serum albumin, a marker of airway epithelial permeability in excess of total protein. We conclude that O 3 exposure in early life induces AHR to nebulized methacholine that is largely dependent on vagal innervation and increased airway epithelial permeability, and to a lesser extent on impaired smooth muscle relaxation.

Su2119 Introduction of Enteric Methanogens Reduces Intestinal Transit and Alters Oral Glucose Tolerance

min functional MRI scan, followed by 10 min acid infusion, and followed by a 6 min functional MRI scan. This protocol was repeated after bilateral vagotomy. For functional imaging we acquired gradient ECHO images on a 9.4T Bruker MRI scanner with the following EPI parameters-matrix 96 x 96,FOV 35mm,TR=2s,TE=18.76 ms, 1 mm slice. All analysis was performed using FSL/FLIRT for image registration and AFNI(NIH) for connectivity analysis. We used seed-based connectivity analysis with placement of seeds within the cingulate cortex (anterior, middle, and retrosplenial). Group comparisons were made using ANOVA and Monte-Carlo simulations for multiple comparison correction. Correlation maps were generated and plotted with a corrected p-value. Results: We found that across all correlation coefficient thresholds, esophageal acid infusion was associated with significantly fewer correlated cortical voxels to the cingulate cortex (Figure 1). We found that when placing seeds within the anterior cingulate cortex, there was statistically significant diminished connectivity with the primary and secondary somatosensory, and primary and secondary motor areas after acid infusion. These alternations were not observed following bilateral cervical vagotomy (Figure 2). The findings of diminished connectivity were also demonstrated when placing seeds in the middle cingulate cortex and retrosplenial cortex. (p-value threshold of 0.02, cluster size of 6 voxels) Conclusions: Esophageal acid stimulation alters functional connectivity of the acid responsive regions of the brain. Esophageal acid induced brain functional connectivity changes are vagally mediated. These findings may play a role in further understanding of acid-related esophageal disorders.

Systemic CCK increases brown adipose tissue sympathetic nerve activity

The neural circuits responsible for governing sympathetic outflow to brown adipose tissue in response to thermoregulatory and febrile stimuli are very well characterized. In contrast we are only beginning to understand the organization of the metabolic regulation of the sympathetic outflow to brown adipose. The current study determines whether cholecystokine (CCK), a gut hormone released postprandially, activates brown adipose tissue sympathetic nerve activity via the vagus nerve. In urethane/chloralose anesthetized rats (n=3), intravenous administration of cholecystokinin (CCK, 0.5μg/kg) increased brown adipose tissue sympathetic nerve activity (BAT SNA: +141 ± 39% control), brown adipose tissue temperature (+0.5 ± 0.1 oC) and expired CO2 (+0.5 ± 0.1%). The CCK‐evoked increases in BAT SNA, BAT temperature and expired CO2 were sustained for at least 40 minutes. Bilateral cervical vagotomy prevented the CCK‐evoked increases in BAT SNA, BAT temperature and expired CO2 (n=2). These results indicate that peripheral CCK acts via vagal afferents to increase metabolism, energy expenditure and thermogenesis in BAT. The neural circuitry mediating this effect and the physiological significance of this system are under investigation.

The effect of heart rate on the heart rate variability response to autonomic interventions

Heart rate variability (HRV), the beat-to-beat variation in either heart rate (HR) or heart period (R-R interval), has become a popular clinical and investigational tool to quantify cardiac autonomic regulation. However, it is not widely appreciated that, due to the inverse curvilinear relationship between HR and R-R interval, HR per se can profoundly influence HRV. It is, therefore, critical to correct HRV for the prevailing HR particularly, as HR changes in response to autonomic neural activation or inhibition. The present study evaluated the effects of HR on the HRV response to autonomic interventions that either increased (submaximal exercise, n = 25 or baroreceptor reflex activation, n = 20) or reduced (pharmacological blockade: β-adrenergic receptor, muscarinic receptor antagonists alone and in combination, n = 25, or bilateral cervical vagotomy, n = 9) autonomic neural activity in a canine model. Both total (RR interval standard deviation, RRSD) and the high frequency (HF) variability (HF, 0.24–1.04 Hz) were determined before and in response to an autonomic intervention. All interventions that reduced or abolished cardiac parasympathetic regulation provoked large reductions in HRV even after HR correction [division by mean RRsec or (mean RRsec)2 for RRSD and HF, respectively] while interventions that reduced HR yielded mixed results. β-adrenergic receptor blockade reduced HRV (RRSD but not HF) while both RRSD and HF increased in response to increases in arterial blood (baroreceptor reflex activation) even after HR correction. These data suggest that the physiological basis for HRV is revealed after correction for prevailing HR and, further, that cardiac parasympathetic activity is responsible for a major portion of the HRV in the dog.

On the pulmonary toxicity of oxygen. 5. Electronic structure and the paramagnetic property of oxygen

Oxygen uptake by the pulmonary circulation is a chemical reaction. The physicochemical attributes of oxygen are critical when studying pulmonary oxygen toxicity. Extent of lung injury depends on the percentage of oxygen in an oxygen–nitrogen mix in polybaric circumstances (Shanklin, 1969). Further change in extent of lesion follows when other gases are used in the inhalant mix instead of nitrogen (Shanklin and Lester, 1972), with oxygen at 21–100% of the mix. Comparative subatmospheric oxygen levels down to 3% in hydrogen, helium, nitrogen, argon, or sulfur hexafluoride, were run with and without ventilatory distress by the Farber (1937) model, bilateral cervical vagotomy (BCV). This yielded coherent results indicating a need to consider molecular characteristics at the atomic level. Molecular mass and size, gas viscosity, and thermal conductivity yielded no obvious correlates to lung injury. Saturation of the outer electron shells of the diluents fit the empiric data, prospectively an interaction between oxygen and nitrogen from their electronegativity and closely approximate molecular mass, size, and shape. The lesion is essentially eliminated at 7% oxygen in nitrogen. At 3% oxygen, the least lesion is found with N2, H2, and SF6, all gases with incomplete outer electron shells, allowing for transient, possibly polarized, covalent bonding with oxygen as the significant minority component in the mix. Argon and helium do not interfere with oxygen. With 3% oxygen in argon without BCV, the experiments ran so long (>70hours) they were terminated once the point had been made. 3% oxygen in argon after BCV yielded a mean survival more than twice that of BCV in air, indicating a remarkable degree of nitrogen interference with oxygen in the respiratory medium of terrestrial animal life. Argon displayed other advantages for the lung compared to nitrogen. Hydrogen, nitrogen, and oxygen are diatomic molecules, a feature which does relate to the extent of lung injury, but only oxygen is paramagnetic.Magnetic effects on lesion formation were tested: [1] with ventilatory distress induced in newborn rabbits, and [2] in young adult female white mice exposed to 100% oxygen without added mechanical distress. A noninvasive model for ventilatory distress, thoracic restraint (TR), with longer mean survivals of 40–50hours, was employed rather than the Farber model. Parallel runs with TR, one subset receiving 100% oxygen in a plastic chamber resting on six strong ring magnets with measured fields up to +1200 gauss, the other plain 100% oxygen, were performed. Both subsets developed moderate metabolic acidosis with average weight losses circa 25%, but over different time courses, 82.89±4.91hours in magnetized oxygen, 55.4% longer than the 53.34±9.82hours in plain oxygen (p<0.001). The longer survival in magnetized oxygen meant extensive lung injury (99.57±0.42% pleural surface, versus 83.86±14.03%), but the rate of lesion formation was 30.89% faster in plain oxygen (1.5722% per hour) than in magnetized oxygen (1.2012% per hour), a difference significant at p<0.001.The effect of oxygen without mechanical ventilatory distress was examined in female adult white mice exposed to oxygen or magnetized oxygen. Similar survivals and weight losses were achieved. The rate of lung lesion formation was different, 1.2617% per hour in plain oxygen, 46.13% faster than 0.8634% per hour in magnetized oxygen. A variable magnetic field, with animals moving and breathing in chambers flooded with oxygen, has both systemic and pulmonary effects which alter the rate of lesion formation due to oxygen toxicity. Paramagnetic oxygen in a magnetic field influences the effect of oxygen toxicity on the lung but at these strengths of field it does not overcome significant mechanical disturbance.

Nucleus ambiguus cholinergic neurons activated by acupuncture: Relation to enkephalin

Acupuncture regulates autonomic function. Our previous studies have shown that electroacupuncture (EA) at the Jianshi–Neiguan acupoints (P5–P6, underlying the median nerve) inhibits central sympathetic outflow and attenuates excitatory cardiovascular reflexes, in part, through an opioid mechanism. It is unknown if EA at these acupoints influences the parasympathetic system. Thus, using c-Fos expression, we examined activation of nucleus ambiguus (NAmb) neurons by EA, their relation to cholinergic (preganglionic parasympathetic) neurons and those containing enkephalin. To enhance detection of cell bodies containing enkephalin, colchicine (90–100μg/kg) was administered into the subarachnoid space of cats 30h prior to EA or sham-operated controls for EA. Following bilateral barodenervation and cervical vagotomy, either EA for 30min at P5–P6 acupoints or control stimulation (needle placement at P5–P6 without stimulation) was applied. While perikarya containing enkephalin were observed in some medullary nuclei (e.g., raphé), only enkephalin-containing neuronal processes were found in the NAmb. Compared to controls (n=4), more c-Fos immunoreactivity, located principally in close proximity to fibers containing enkephalin was noted in the NAmb of EA-treated cats (n=5; P<0.01). Moreover, neurons double-labeled with c-Fos and choline acetyltransferase in the NAmb were identified in EA-treated, but not control animals. These data demonstrate for the first time that EA activates preganglionic parasympathetic neurons in the NAmb. Because of their close proximity, these EA-activated neurons likely interact with nerve fibers containing enkephalin. These results suggest that EA at the P5–P6 acupoints has the potential to influence parasympathetic outflow and cardiovascular function, likely through an enkephalinergic mechanism.

On the pulmonary toxicity of oxygen. 4. The thyroid arena

Normally developed thyroid function is critical to the transition from fetal to neonatal life with the onset of independent thermoregulation, the most conspicuous of the many ways in which thyroid secretions act throughout the body. A role for thyroid secretions in growth and maturation of the lungs as part of the preparation for the onset of breathing has been recognized for some time but how this contributes to tissue and cell processes and defenses under the duress of respiratory distress has not been well examined. Extensive archival autopsy material was searched for thyroid and adrenal weights, first by gestational age, and then for changes during the first hours after birth as ratios to body weight. After a gestational age of 22weeks the fetal thyroid and adrenal glands at autopsy in those with hyaline membrane disease are persistently half the size of those in “normal” infants dying with other disorders. When the thyroid is examined shortly after birth it reveals a post natal loss of mass per body weight of similar orders of magnitude which does not occur in the control group. A clinical sample of premature infants with (12) and without (14) hyaline membrane disease was tested for T4, TSH, TBG, and total serum protein. The results also demonstrate a special subset with lower birth weights at the same gestational age, and lower serum T4 and total serum protein. Ventilatory distress in newborn rabbits was induced by bilateral cervical vagotomy at 24h post natal following earlier injection of thyroxine (T4) or thyroid stimulating hormone (TSH) and comparisons were made with untreated animals and by dose. Early life thyroidectomy was performed followed by exposure to either air or 100% oxygen. A final experiment in air was vagotomy after thyroidectomy. Composite analysis of these methods indicates that thyroid factors are both operative and important in the newborn animal with ventilatory distress. This work and the archival data indicate those infants destined to develop hyaline membrane disease through respiratory distress are a distinct developmental and clinical subset with the point of departure from otherwise normal development and maturation in the second or early third trimester. This interval is known to be a period of marked variation in the overview indicators of fetal progress through gestational time. The initiating factor or circumstance which then separates this special subset from normal future development is placed by these observations firmly into the period when human fetal TSH dramatically rises 7-fold (17.5–25.5weeks) followed by a lesser 3 to 4-fold increase in T4 which is extended into the early third trimester. The earlier part of this interval is characterized by the thyrotrophic action of chorionic gonadotropin (hCG). The possibility that abnormalities in the intrauterine environment secondary to maternal infection play a role within this time frame is indicated by the demonstration that interleukin-2 (IL-2) induces an anterior pituitary release of TSH. Since IL-2 has this property and is not an acute phase cytokine, some form of chronic infection or an immunopathic process seems more likely as a possible active factor in pathogenesis.

Effect of esophagus distention on urinary bladder function in rats

Micturition process is a spinobulbospinal reflex that is affected by the viscero-visceral interactions due to convergent inputs into spinal and/or supraspinal centers controlling that reflex. Although interaction between bladder and other pelvic organs, such as colon, are well studied, the viscero-visceral interaction between urinary bladder and internal organs in other regions are rarely studied. In the present study, continuous filling cystometry recordings, in male rats, were used to investigate the effects of mechanical stimulation of distal-esophagus (distention), as well as, electrical stimulation of abdominal branches of the vagus nerve on urinary bladder micturition cycles. Distal esophagus distention and electrical stimulation of the vagus nerve significantly increased the micturition frequency through decreasing the time of the storage phase of the micturition cycle. However, bilateral cervical vagotomy eliminated the effects of distal esophagus distention and electrical stimulation of vagus nerve on micturition cycles. The results of this study indicate that there is a viscero-visceral interaction between esophagus and urinary bladder, which is mediated through vagal afferents. Understanding the properties of the viscero-visceral interactions affecting the urinary bladder will help in the diagnosis and management of micturition problems.

Variability in the muscle composition of rat esophagus and neural pathway of lower esophageal sphincter relaxation

Several studies from our laboratory show that axial stretch of the lower esophageal sphincter (LES) in an oral direction causes neurally mediated LES relaxation. Under physiological conditions, axial stretch of the LES is caused by longitudinal muscle contraction (LMC) of the esophagus. Because longitudinal muscle is composed of skeletal muscle in mice, vagal-induced LMC and LES relaxation are both blocked by pancuronium. We conducted studies in rats (thought to have skeletal muscle esophagus) to determine if vagus nerve-mediated LES relaxation is also blocked by pancuronium. LMC-mediated axial stretch on the LES was monitored using piezoelectric crystals. LES and esophageal pressures were monitored with a 2.5-Fr solid-state pressure transducer catheter. Following bilateral cervical vagotomy, the vagus nerve was stimulated electrically. LES, along with the esophagus, was harvested after in vivo experiments and immunostained for smooth muscle (smooth muscle α-actin) and skeletal muscle (fast myosin heavy chain). Vagus nerve-stimulated LES relaxation and esophageal LMC were reduced in a dose-dependent fashion and completely abolished by pancuronium (96 μg/kg) in six rats (group 1). On the other hand, in seven rats, LES relaxation and LMC were only blocked completely by a combination of pancuronium (group 2) and hexamethonium. Immunostaining revealed that the longitudinal muscle layer was composed of predominantly skeletal muscle in the group 1 rats. On the other hand, the longitudinal muscle layer of group 2 rats contained a significant amount of smooth muscle (P < 0.05). Our study shows tight coupling between axial stretch on the LES and relaxation of the LES, which suggests a cause and effect relationship between the two. We propose that the vagus nerve fibers that cause LMC induce LES relaxation through the stretch-sensitive activation of inhibitory motor neurons.

Melanocortin 4 Receptor Activation Protects Against Testicular Ischemia-Reperfusion Injury by Triggering the Cholinergic Antiinflammatory Pathway

Melanocortins (MC) trigger a vagus nerve-mediated cholinergic-antiinflammatory pathway projecting to the testis. We tested whether pharmacological activation of brain MC receptors might protect the testis from the damage induced by ischemia-reperfusion. Adult male rats were subjected to 1-h testicular ischemia, followed by 24-h reperfusion [testicular ischemia-reperfusion (TI/R)]. Before TI/R, groups of animals were subjected to bilateral cervical vagotomy, or pretreated with the nicotinic acetylcholine receptor antagonist chlorisondamine or the selective MC(4) receptor antagonist HS024. Immediately after reperfusion, rats were ip treated with saline or the MC analog [Nle(4),D-Phe(7)]α-melanocyte-stimulating hormone (NDP-α-MSH) (340 μg/kg). We evaluated testicular IL-6 and TNF-α by Western blot analysis and organ damage by light microscopy. Some experimental groups were prepared for neural efferent activity recording along the vagus nerve starting 30 min after treatment with NDP-α-MSH or saline, and for a 30-min period. Additional groups of TI/R rats were treated for 30 d with saline, NDP-α-MSH, chlorisondamine plus NDP-α-MSH, or HS024 plus NDP-α-MSH to evaluate spermatogenesis, organ damage, and the apoptosis machinery. After a 24-h reperfusion, in TI/R saline-treated rats, there was an increase in IL-6 and TNF-α expression and a marked damage in both testes. NDP-α-MSH inhibited IL-6 and TNF-α expression, decreased histological damage, and increased neural efferent activity. Furthermore, NDP-α-MSH administration for 30 d greatly improved spermatogenesis, reduced organ damage, and inhibited apoptosis. All positive NDP-α-MSH effects were abrogated by vagotomy, chlorisondamine, or HS024. Our data suggest that selective MC(4) receptor agonists might be therapeutic candidates for the management of testicular torsion.