Effects of feeding diets based on transgenic soybean meal and soybean hulls to dairy cows on production measures and sensory quality of milk.

This experiment was conducted to determine whether feeding meal and hulls derived from genetically modified soybeans to dairy cows affected production measures and sensory qualities of milk. The soybeans were genetically modified (Event DAS-444Ø6-6) to be resistant to multiple herbicides. Twenty-six Holstein cows (13/treatment) were fed a diet that contained meal and hulls derived from transgenic soybeans or a diet that contained meal and hulls from a nontransgenic near-isoline variety. Soybean products comprised approximately 21% of the diet dry matter, and diets were formulated to be nearly identical in crude protein, neutral detergent fiber, energy, and minerals and vitamins. The experimental design was a replicated 2×2 Latin square with a 28-d feeding period. Dry matter intake (21.3 vs. 21.4kg/d), milk yield (29.3 vs. 29.4kg/d), milk fat (3.70 vs. 3.68%), and milk protein (3.10 vs. 3.12%) did not differ between cows fed control or transgenic soybean products, respectively. Milk fatty acid profile was virtually identical between treatments. Somatic cell count was significantly lower for cows fed transgenic soybean products, but the difference was biologically trivial. Milk was collected from all cows in period 1 on d 0 (before treatment), 14, and 28 for sensory evaluation. On samples from all days (including d 0) judges could discriminate between treatments for perceived appearance of the milk. The presence of this difference at d 0 indicated that it was likely not a treatment effect but rather an initial bias in the cow population. No treatment differences were found for preference or acceptance of the milk. Overall, feeding soybean meal and hulls derived from this genetically modified soybean had essentially no effects on production or milk acceptance when fed to dairy cows.

Activation of brain stem neurons by irritant chemical stimulation of the throat assessed by c-fos immunohistochemistry.

The method of c-fos immunohistochemistry was used to identify the brain stem distribution of neurons activated following irritant chemical stimulation of the laryngopharyngeal mucosa. In pentobarbital-anesthetized rats, either water (control), nicotine (600 mM, 1 ml) or capsaicin (330 microM, 1 ml) was applied to the pharynx via a cannula placed posterior to the soft palate. Following nicotine and capsaicin, there was a significant increase in fos-like immunoreactivity (FLI) compared with controls in the following areas: nucleus of the solitary tract from the level of the pyramidal decussation caudally to the level of the area postrema rostrally; dorsomedial aspect of trigeminal subnucleus caudalis (Vc); and paratrigeminal islands interspersed in the spinal trigeminal tract. There was significantly more FLI in Vc and paratrigeminal nuclei following capsaicin than following nicotine, while the reverse was true for NTS. In addition, there was a significant increase in FLI in area postrema and the ventrolateral medullary region dorsal to the lateral reticular nucleus following nicotine but not capsaicin. The distributions of FLI in NTS, area postrema, Vc, and paratrigeminal nuclei are consistent with prior anatomical tract-tracing studies and suggest roles for these brain stem regions in mediating sensory and reflex responses to irritant chemical stimulation of the upper respiratory mucosa.

Analgesia induced by chronic nicotine infusion in rats: differences by gender and pain test.

RATIONALE: Acute administration of nicotine induces analgesia with subsequent development of tolerance. In human studies, females are less sensitive to the analgesic effects of nicotine than males. Few previous animal studies have investigated analgesic effects of chronic nicotine administration or addressed gender differences. OBJECTIVES: To investigate whether chronic administration of nicotine induces analgesia in male and female rats as assessed by a battery of standard pain assays, if tolerance develops, and if hyperalgesia occurs following cessation of nicotine. METHODS: Nicotine (free base; 6 mg/kg/day i.v.) or saline was administered for 2 weeks via implanted osmotic pumps. Pain behavior was assessed before, during, and for 3 weeks after nicotine infusion by measuring tail flick latency, hot-plate latency, and thermal paw withdrawal latency. The paw-withdrawal threshold to non-noxious mechanical stimuli was also measured. Effects of nicotine infusion, gender, and time were assessed by three-way analyses of variance. RESULTS: Both male and female rats exhibited a comparable degree of analgesia in the hot-plate test with development of tolerance during the 2-week infusion period. Males, but not females, showed analgesia in the tail flick test. Analgesia was not observed for thermally evoked paw withdrawal in either males or females, nor did nicotine affect non-noxious mechanically evoked paw withdrawals. Males and females showed cessation of weight gain during the first week of nicotine infusion. CONCLUSIONS: Chronic nicotine-induced analgesia was confirmed in both male and female rats as assessed using the hot-plate test which reflects integrated pain behavior. Males, but not females, exhibited analgesia in a nociceptive withdrawal reflex test (tail flick), indicating that nicotine-induced analgesia may depend on both the type of pain test and gender. The lack of nicotine-induced analgesia assessed by the tail flick reflex test in female rats is consistent with recent human studies showing that nicotine reduces pain elicited by brief noxious cutaneous stimulation in male but not female subjects.

The oral sensation of carbonated water: cross-desensitization by capsaicin and potentiation by amiloride.

The oral sensation elicited by carbonated water is reduced by capsaicin and by blockers of carbonic anhydrase. We have investigated the temporal profile of this sensation and its cross-desensitization by capsaicin. We additionally tested if the sensation is influenced by amiloride. Following pretreatment of half of the dorsal tongue with 33 p.p.m. capsaicin, carbonated water was flowed over the tongue bilaterally for 5, 15 or 60 s. Subjects then performed a two-alternative forced choice test by indicating which side of the tongue had a stronger sensation and separately rated the sensory intensity on each side. Capsaicin significantly reduced the intensity of sensation elicited by carbonated water, consistent with cross-desensitization. This effect was weaker at 60 s because of a significant decline (desensitization) in ratings of the intensity of carbonated water on both sides of the tongue. Pretreatment with amiloride resulted in a small but significant increase in the intensity of the sensation elicited by the 15 s carbonated water stimulus, suggesting an amiloride-sensitive transduction mechanism.

An animal model to assess aversion to intra-oral capsaicin: increased threshold in mice lacking substance p.

Despite the widespread consumption of products containing chemicals that irritate the oral mucosa, little is known about the underlying neural mechanisms nor is there a corresponding animal model of oral irritation. We have developed a rodent model to assess aversion to capsaicin in drinking water, using a paired preference paradigm. This method was used to test the hypothesis that the neuromodulator substance P (SP) plays a role in the detection of intra-oral capsaicin. 'Knockout' (KO) mice completely lacking SP and neurokinin A due to a disruption of the preprotachykinin A gene and a matched population of wild-type (WT) mice had free access to two drinking bottles, one containing water and the other capsaicin at various concentrations. Both KO and WT mice showed a concentration-dependent aversion to capsaicin. KO mice consumed significantly more capsaicin than WT at a single near threshold (1.65 microM) concentration, indicating that SP plays a limited role in the detection and rejection of oral irritants.

Sensitization, desensitization and stimulus-induced recovery of trigeminal neuronal responses to oral capsaicin and nicotine.

Repeated application of capsaicin at a 1-min interstimulus interval (ISI) to the tongue induces a progressively increasing irritant sensation (sensitization), followed after a rest period by reduced sensitivity to further capsaicin (desensitization). Sequential reapplication of capsaicin induces irritation that eventually increases to initial levels: stimulus-induced recovery (SIR). In contrast, repeated application of nicotine elicits a declining irritant sensation across trials. To investigate possible neural correlates of these phenomena, we recorded from single units in superficial laminae of the dorsomedial trigeminal subnucleus caudalis (Vc) that responded to noxious thermal (54 degrees C) and chemical (1 M pentanoic acid) stimulation of the tongue of anesthetized rats. We then recorded responses to either capsaicin (330 microM) or nicotine (0.6 M), delivered either once, repeatedly at 1-min ISI, or continually by constant flow. After the initial capsaicin application and a rest period, the capsaicin was reapplied in the identical manner to test for SIR. The mean response of 14 Vc units to sequential application of pentanoic acid did not vary significantly across trials, indicating lack of tachyphylaxis or sensitization. The averaged response of 11 Vc units to repeated capsaicin increased significantly across the first eight trials and then plateaued. Following the rest period, spontaneous firing had returned to the precapsaicin level. With capsaicin reapplication, the averaged response increased again after a significant delay (due to desensitization), but did not reattain the peak firing rate achieved in the initial series (partial SIR). Constant-flow application of capsaicin induced an identical sensitization followed by nearly complete SIR. A single application of capsaicin induced a significant rise in firing in eight other units, but the rate of rise and maximal firing rate were both much lower compared with repetitive or constant-flow capsaicin. When capsaicin was reapplied once after the rest period, there was no change in firing rate indicating absence of SIR. These results indicate that maintenance of the capsaicin concentration induces a progressive increase in neuronal response that parallels sensitization. With recurrent capsaicin application, desensitization can be overcome to result in a delayed recovery of Vc responses similar to SIR. In contrast, the averaged response of 17 Vc units to repeated or constant-flow application of nicotine increased only over the first 3 min, and then decreased to spontaneous levels even as nicotine was still being applied. These results are consistent with the decrease in the perceived irritation elicited by sequential application of nicotine in humans.

Role of neuronal nicotinic-acetylcholine receptors in the activation of neurons in trigeminal subnucleus caudalis by nicotine delivered to the oral mucosa.

To characterize the role of neuronal nicotinic acetylcholine receptors (nAChRs) in oral irritation and pain, we employed the method of c-fos immunohistochemistry to map the locations and numbers of brainstem neurons that express the immediate-early gene, c-fos, after application of nicotine to the tongue, either alone or after pretreatment with cholinergic antagonists. Groups of anesthetized rats received the following chemicals delivered bilaterally to the dorsal tongue: (1) 0.9% NaCl followed by nicotine (1%, 61 mM), (2) the nAChR antagonist, mecamylamine 0.1% (= 4.9 mM) followed by nicotine, (3) the muscarinic antagonist atropine (0.1% 1.46 mM) followed by nicotine, (4) atropine (1%, 14.6 mM) followed by nicotine, (5) 0.9% NaCl as a control, and (6) unstimulated controls. Two hours later, animals were perfused with phosphate-buffered saline followed by 4% paraformaldehyde through the aorta. Post-fixed brainstems were cut in 50-micron frozen sections and immunohistochemically processed for fos-like immunoreactivity (FLI). Following application of nicotine, there were significant increases in FLI compared with saline-treated controls in dorsomedial and ventrolateral aspects of the trigeminal caudalis. Pretreatment with either mecamylamine or the high (1%) concentration of atropine significantly reduced nicotine-evoked FLI in these areas, while pretreatment with the low (0.1%) atropine concentration did not significantly affect FLI. These results are consistent with the idea that nicotine activates nAChRs residing on lingual nociceptive fibers, which, in turn, excite neurons in trigeminal caudalis.

Psychophysical and neurobiological evidence that the oral sensation elicited by carbonated water is of chemogenic origin.

The sensation produced by carbonated beverages has been attributed to chemical excitation of nociceptors in the oral cavity via the conversion of CO(2) to carbonic acid in a reaction catalyzed by carbonic anhydrase. In separate studies, we tested if the carbonic anyhdrase blocker, acetazolamide, reduced either the intensity of sensation in humans or c-fos expression by trigeminal neurons in rats, evoked by application of carbonated water to the tongue. In the psychophysical experiment, one-half of the dorsal tongue was pretreated with acetazolamide (1 or 2%), after which the tongue was exposed bilaterally to carbonated water. In a two-alternative forced-choice paradigm, subjects chose which side of the tongue yielded a stronger sensation and additionally rated the magnitude of sensation on each side. Pretreatment with acetazolamide reduced the magnitude of sensation elicited by carbonated water in a concentration-dependent manner, since a significant majority of subjects chose the untreated side of the tongue as having a stronger sensation and assigned significantly higher intensity ratings to that side. Acetazolamide did not affect the irritant sensation from citric acid, while capsaicin pretreatment reduced both the sensation elicited by carbonated water and the irritation induced by citric acid application. In a separate experiment using rats, delivery of carbonated water to the tongue significantly increased the number of cells expressing c-fos-like immunoreactivity in the dorsomedial trigeminal nucleus caudalis (versus saline controls); this was significantly reduced by pretreatment with acetazolamide. Our results support the hypothesis that carbonated water activates lingual nociceptors via conversion of CO(2) to carbonic acid; the nociceptors in turn excite trigeminal neurons involved in signaling oral irritation.

Does the formation of lipopolysaccharide tolerance require intact vagal innervation of the liver?

The study was designed to test whether intact vagal innervation of the liver is required for the formation of tolerance to lipopolysaccharide (LPS). Wistar rats were subjected to either denervation of the liver (transection of the hepatic and both celiac branches of the abdominal vagus) or sham surgery. Two weeks later, each rat had an osmotic pump implanted subcutaneously. The pump was filled with either a suspension of Escherichia coli LPS (18 mg/ml) in saline or saline alone. Via a catheter, the pump delivered its content into the right jugular vein at a rate of approximately 0.72 microl/kg/h (approximately 13 microg/kg/h of LPS) over 28 d. On day 25 of the infusion, each animal had another catheter implanted into the left jugular vein. Three days later, each rat was injected with a lethal bolus dose of LPS (15 mg/kg) and had its colonic temperature recorded. The saline-infused sham-operated rats responded to the bolus injection of LPS with hypothermia followed by a fever (mean response magnitude 1.0+/-0.2 degrees C); 91% of the animals died within 24 h. The LPS-primed shams developed marked tolerance: When challenged with a lethal dose of LPS, they exhibited a significantly smaller thermal response (magnitude 0.5 +/- 0.2 degrees C) and none died. No group of the vagotomized animals, whether LPS- or saline-primed, became tolerant: Both groups exhibited similar hypothermic responses to the bolus LPS injection and a substantial mortality rate (40 and 100%, respectively). The study shows that prolonged infusion of low doses of LPS leads to the formation of tolerance and that vagal denervation of the liver by hepato-celiac vagotomy suppresses this process. The mechanisms of vagal control of the formation of LPS tolerance remain speculative.

Multiple neural mechanisms of fever.

In rats, fevers induced by moderate-to-high doses of intravenous lipopolysaccharide consist of three phases (phases 1, 2 and 3) with body temperature peaks at approximately 1, 2, and 5 h postinjection, respectively. In this study, the effects of bilateral truncal subdiaphragmatic vagotomy and intraperitoneal capsaicin desensitization on febrile phases 1-3 were assessed in adult Wistar rats. Surgical vagotomy was performed approximately 30 d before the experiment; this procedure interrupts both afferent and efferent vagal fibers. Capsaicin was administered intraperitoneally in two consecutive injections (2 and 3 mg/kg, 3 h apart) 1 week prior to the experiment; this procedure desensitizes afferent fibers, primarily within the abdominal cavity, and does not lead to the known thermal effects of systemic capsaicin desensitization. At a neutral ambient temperature, the rats were given Escherichia coli lipopolysaccharide (10 microg/kg) through a preimplanted jugular catheter, and their colonic temperature wes measured by thermocouples for 7 h. The control rats exhibited the typical triphasic febrile responses. Confirming our earlier studies, subdiaphragmatic vagotomy did not affect phases 1 and 2; it did, however, result in a 2.5-fold reduction of phase 3. Capsaicin desensitization modified the febrile response differently: phases 2 and 3 were unaffected, but phase 1 disappeared. We suggest that neural afferent fibers (nonvagal but perhaps vagal as well) play an important role in the early febrile response (phase 1) by transducing peripheral pyrogenic signals to the brain. We also suggest that vagal efferent fibers are likely to participate in the later febrile response (phase 3) via an unknown mechanism.

Vagotomy does not affect thermal responsiveness to intrabrain prostaglandin E2 and cholecystokinin octapeptide.

Subdiaphragmatic vagotomy has been repeatedly shown to attenuate the febrile response to peripherally injected pyrogens. In the present study, we investigated whether vagotomy-induced attenuation of febrile responsiveness reflects a decreased sensitivity of the brain to central fever mediators, prostaglandin E2 (PGE2) and cholecystokinin octapeptide (CCK-8). Male rats were subjected to subdiaphragmatic vagotomy (or sham surgery) on day 0 and had a cannula implanted into the lateral cerebral ventricle on day 24. On day 30-36, the thermal responsiveness of the rats to PGE2 or CCK-8 was tested. Each animal was injected in the ventricle with either PGE2 (0, 10, 100, or 500 ng) in pyrogen-free saline with 0.5% ethanol (5 microl) or CCK-8 (0 or 1.6 microg) in artificial cerebro-spinal fluid (5 microl). While the 0-dose of either PGE2 or CCK-8 (vehicle alone) induced no thermal response, all the higher doses of either agent caused a body temperature rise preceded by tail skin vasoconstriction. The vagotomized rats did not respond differently than their sham-operated counterparts to any dose of either drug. It is concluded that subdiaphragmatic vagotomy does not change the rat's thermal responsiveness to intrabrain PGE(2) and CCK-8.

Neurobiological and psychophysical mechanisms underlying the oral sensation produced by carbonated water.

Carbonated drinks elicit a sensation that is highly sought after, yet the underlying neural mechanisms are ill-defined. We hypothesize that CO(2) is converted via carbonic anhydrase into carbonic acid, which excites lingual nociceptors that project to the trigeminal nuclei. We investigated this hypothesis using three methodological approaches. Electrophysiological methods were used to record responses of single units located in superficial laminae of the dorsomedial aspect of trigeminal subnucleus caudalis (Vc) evoked by lingual application of carbonated water in anesthetized rats. After pretreatment of the tongue with the carbonic anhydrase inhibitor dorzolamide, neuronal responses to carbonated water were significantly attenuated, followed by recovery. Using c-Fos immunohistochemistry, we investigated the distribution of brainstem neurons activated by intraoral carbonated water. Fos-like immunoreactivity (FLI) was significantly higher in the superficial laminae of dorsomedial and ventrolateral Vc in animals treated with carbonated water versus controls. Dorzolamide pretreatment significantly reduced FLI in dorsomedial Vc. We also examined the sensation elicited by carbonated water in human psychophysical studies. When one side of the tongue was pretreated with dorzolamide, followed by bilateral application of carbonated water, a significant majority of subjects chose the untreated side as having a stronger sensation and assigned significantly higher intensity ratings to that side. Dorzolamide did not reduce irritation elicited by pentanoic acid. The present data support the hypothesis that carbonated water excites lingual nociceptors via a carbonic anhydrase-dependent process, in turn exciting neurons in Vc that are presumably involved in signaling oral irritant sensations.

Signaling the brain in systemic inflammation: which vagal branch is involved in fever genesis?

Recent evidence has suggested a role of abdominal vagal afferents in the pathogenesis of the febrile response. The abdominal vagus consists of five main branches (viz., the anterior and posterior celiac branches, anterior and posterior gastric branches, and hepatic branch). The branch responsible for transducing a pyrogenic signal from the periphery to the brain has not as yet been identified. In the present study, we address this issue by testing the febrile responsiveness of male Wistar rats subjected to one of four selective vagotomies: celiac (CBV), gastric (GBV), hepatic (HBV), or sham (SV). In the case of CBV, GBV, and HBV, only the particular vagal branch(es) was cut; for SV, all branches were left intact. After the postsurgical recovery (26-29 days), the rats had a catheter implanted into the jugular vein. On days 29-32, their colonic temperature (Tc) responses to a low dose (1 microg/kg) of Escherichia coli lipopolysaccharide (LPS) were studied. Three days later, the animals were subjected to a 24-h food and water deprivation, and the effectiveness of the four vagotomies to induce gastric food retention, pancreatic hypertrophy, and impairment of the portorenal osmotic reflex was assessed by weighing the stomach and pancreas and measuring the specific gravity of bladder urine, respectively. Stomach mass, pancreas mass, and urine density successfully separated the four experimental groups into four distinct clusters, thus confirming that each type of vagotomy had a different effect on the indexes measured. The Tc responses of SV, CBV, and GBV rats to LPS did not differ and were characterized by a latency of approximately 40 min and a maximal rise of 0.7 +/- 0.1, 0.6 +/- 0.1, and 0.9 +/- 0.2 degrees C, respectively. The fever response of the HBV rats was different; practically no Tc rise occurred (0.1 +/- 0.2 degrees C). The HBV appeared to be the only selective abdominal vagotomy affecting the febrile responsiveness. We conclude, therefore, that the hepatic vagus plays an important role in the transduction of a pyrogenic signal from the periphery to the brain.

"Biphasic" fevers often consist of more than two phases.

This paper disproves the common belief that all doses of lipopolysaccharide (LPS) that are commonly referred to as biphasic fever inducing (>/=2 microg/kg) cause truly biphasic responses. A catheter was implanted into the right jugular vein of several strains of adult male rats, and the animals were habituated to the experimental conditions. At an ambient temperature of 30.0 degrees C, loosely restrained animals were injected with a 10 microg/kg dose of LPS (various preparations), and their colonic (Tc) and tail skin temperatures were monitored (from >/=1 h before to >/=7 h after the injection). The results are presented as time graphs and phase-plane plots; in the latter case the rate of change of Tc is plotted against Tc. In experiment 1 the intravenous injection of LPS (from Escherichia coli 0111:B4, phenol extract) into the rats (Bkl:Wistar) induced a triphasic febrile response, as is obvious from time graphs of Tc (3 peaks), time graphs of effector activity (3 waves of tail skin vasoconstriction), and phase-plane plots (3 complete loops); the injection of saline (control) induced no Tc changes. We analyzed whether the triphasic pattern was due to some peculiarities of the experimental design, i.e., the pyrogen preparation used (experiment 2) or the rat strain tested (experiment 3) or whether this pattern reflects a more general law. In experiment 2 we used the same (phenol) preparation of different LPS (from Shigella flexneri 1A and Salmonella typhosa) and a different preparation (TCA extract) of the same LPS (E. coli). Regardless of the LPS used, rats of the Bkl:Wistar strain responded to the 10 microg/kg dose with the triphasic fever. In experiment 3, rats of other strains [Bkl:Sprague-Dawley and Sim:(LE)fBR(Black-hooded)] were tested. Again, all animals responded to the 10 microg/kg dose of E. coli LPS (phenol extract) with the triphasic fever. Because all fevers caused by four different LPS preparations in three rat strains were triphasic, the triphasic pattern is likely to constitute an intrinsic characteristic of the febrile response.

Methodology of fever research: why are polyphasic fevers often thought to be biphasic?

This study explains why the recently described triphasic lipopolysaccharide (LPS) fevers have been repeatedly mistaken for biphasic fevers. Experiments were performed in loosely restrained male Wistar rats with a catheter implanted into the right jugular vein. Each animal was injected with Escherichia coli LPS, and its colonic (Tc) and tail skin temperatures were monitored. The results are presented as time graphs and phase-plane plots; in the latter case the rate of change of Tc is plotted against Tc. At an ambient temperature (Ta) of 30.0 degrees C, the response to the 10 microg/kg dose of LPS was triphasic, as is obvious from time graphs of Tc (3 peaks), time graphs of effector activity (3 waves of tail skin vasoconstriction), and phase-plane plots (3 complete loops). When the Ta was below neutral (22.0 degrees C) or the LPS dose was higher (100 or 1,000 microg/kg), the time graph of Tc did not allow for the reliable detection of all three febrile phases, but the phase-plane plot and time graph of effector activity clearly revealed the triphasic pattern. In a separate experiment, LPS (10 microg/kg) or saline was injected via one of two different procedures: in the first group the injection was performed through the jugular catheter, from outside the experimental chamber; in the second group the same nonstressing injection was combined with opening the chamber and pricking the animal in its lower abdomen with a needle. In the first group the febrile response was obviously triphasic, and none of the phases was due to the procedure of injection per se (injection of saline did not affect Tc). In the second group the fever similarly consisted of three Tc rises, but it might have been readily mistaken for biphasic because the first rise was indistinguishable from stress hyperthermia occurring in the saline-injected (and needle-pricked) controls. We conclude that several methodological factors (dose of LPS, procedure of its injection, and Ta) have contributed, although each in a different way, to the common misbelief that there are only two febrile phases.

Cold defense mechanisms in vagotomized rats.

Subdiaphragmatically vagotomized rats cannot mount a febrile response to pyrogens and are believed to have severe thermoregulatory deficiencies. We addressed the issue of thermoeffector competence of vagotomized rats by asking three questions. In Expt. 1 we asked, can vagotomized rats readily recruit tail skin vasoconstriction in the course of a moderate cold exposure? In Expt. 2 the question was, can brown adipose tissue (BAT) thermogenesis readily be activated in vagotomized rats (e.g., in response to a tail pinch)? In Expt. 3, we investigated the question: can vagotomized rats elevate their body temperature in response to ephedrine (a drug of high hyperthermizing potential) to the same extent as sham-operated controls? Rats were vagotomized or sham operated and implanted with a catheter into the jugular vein and a thermocouple into the interscapular BAT. To prevent the common complications of vagotomy, special perioperative care was given. During experiments, colonic, tail skin, and BAT temperatures (Tc, Tsk, and TBAT, respectively) were measured. The vagotomized animals were well nourished and had a body mass (325 +/- 6 g) similar to that of the controls (338 +/- 6 g). In Expt. 1, in response to external cooling (15 degrees C, 1 h), the vagotomized (n = 30) and sham-operated (n = 31) rats recruited tail skin vasoconstriction at close values of both Tc (37.84 +/- 0.08 and 37.97 +/- 0.07 degrees C) and Tsk (33.16 +/- 0.17 and 33.18 +/- 0.18 degrees C, respectively). In Expt. 2, tail pinch-associated stress in vagotomized rats resulted in a sharp rise in the TBAT-Tc gradient by 0.3-1.0 degree C. In Expt. 3, ephedrine administered intravenously (whether in a 5 or 35 mg/kg dose) evoked similar hyperthermic responses in the vagotomized and sham-operated rats: a moderate (approximately 2.5 degrees C) Tc rise in the low dose and a "supramaximal" (approximately 5.0 degrees C) rise in the high dose. In sum, the answer to all three questions asked is yes. Vagotomized rats, at least when well nourished, exhibit no signs of thermoeffector deficiency. It is, therefore, not effector incompetence but rather vagal deafferentation per se that can explain the febrile irresponsiveness of vagotomized rats.

Febrile responsiveness of vagotomized rats is suppressed even in the absence of malnutrition.

The repeatedly observed attenuation of fever in vagotomized rats has been accepted as evidence of an essential role of vagal afferents in the transduction of pyrogenic signals from the periphery to the brain. If, however, the general condition of a vagotomized animal is poor (the usual case) and accompanied by malnutrition and body mass loss (common complications of vagotomy), the febrile responsiveness can be suppressed not because of the lack of vagal afferentation, but rather secondarily to a malnutrition-associated thermogenic incompetence. In the present study, we addressed this dilemma. Male Wistar rats were subjected to subdiaphragmatic vagotomy (or sham surgery) and, 24 days later, catheterized in the jugular vein. Postsurgically, the rats were closely watched and fed highly palatable food. Their febrile responsiveness [colonic (Tc) and tail skin (Tsk) temperature responses] to Escherichia coli lipopolysaccharide (LPS: 1 microgram/kg i.v.) was tested on day 27 postvagotomy. To verify the completeness of vagotomy, each rat was food deprived for 24 h and then euthanized; its stomach's evacuatory function was assessed by weighing the organ. One month postsurgery, both food consumption and body mass of the vagotomized rats (33 +/- 2 g/day and 313 +/- 4 g, respectively) were similar to the control values (30 +/- 1 g/day and 315 +/- 8 g). In the sham rats, LPS induced a monophasic Tc rise of 0.5 +/- 0.3 degree C at 70 min postinjection (peak), preceded by a fall in Tsk. Neither this Tsk fall (tail skin vasoconstriction) nor the resultant fever occurred in the vagotomized rats; at 70 min, Tc change was -0.1 +/-0.1 degree C. The gastric mass (4.1 +/- 0.5 g in the vagotomized vs. 1.8 +/- 0.1 g in sham rats) indicated the effectiveness of vagotomy. In sum, although the vagotomy-associated malnutrition was successfully prevented with special perioperative care, the vagotomized animals still did not respond to LPS with fever. Malnutrition is, therefore, unlikely to constitute the main reason of the febrile irresponsiveness of vagotomized rats.

The vagus nerve in the thermoregulatory response to systemic inflammation.

Experimentally, systemic inflammation induced by a bolus intravenous injection of lipopolysaccharide (LPS) may be accompanied by three different thermoregulatory responses: monophasic fever (the typical response to low doses of LPS), biphasic fever (medium doses), and hypothermia (high doses). In our recent study [Romanovsky, A. A., V. A. Kulchitsky, C. T. Simons, N. Sugimoto, and M. Székely. Am. J. Physiol. (Regulatory Integrative Comp. Physiol.). In press], monophasic fever did not occur in subdiaphragmatically vagotomized rats. In the present work, we asked whether vagotomy affects the two other types of thermoregulatory response. Adult Wistar rats were vagotomized (or sham operated) and had an intravenous catheter implanted. On day 28 postvagotomy, the thermal responses to the intravenous injection of Escherichia coli LPS (0, 1, 10, 100, or 1,000 micrograms/kg) were tested in either a neutral (30 degrees C) or slightly cool (25 degrees C) environment. Three major results were obtained. 1) In the sham-operated rats, the 1 microgram/kg dose of LPS caused at 30 degrees C a monophasic fever with a maximal colonic temperature (Tc) rise of approximately 0.6 degree C; this response was abated (no Tc changes) in the vagotomized rats. 2) At 30 degrees C, all responses to higher doses of LPS (10-1,000 micrograms/kg) were represented by biphasic fevers (the higher the dose, the less pronounced the first and the more pronounced the second phase was); none of these biphasic fevers was altered in the vagotomized animals. 3) In response to the 1,000 micrograms/kg dose at 25 degrees C, hypothermia occurred: Tc changed by -0.5 +/- 0.1 degree C (nadir); this hypothermia was exaggerated (-1.1 +/- 0.1 degrees C) in the vagotomized rats. It is concluded that vagal afferentation may be important in the mediation of the response to minor amounts of circulating LPS, whereas the response to larger amounts is brought about mostly (if not exclusively) by nonvagal mechanisms. This difference may be explained by the dose-dependent mechanisms of the processing of exogenous pyrogens. Vagotomized animals also appear to be more sensitive to the hypothermizing action of LPS in a cool environment; the mechanisms of this phenomenon remain speculative.