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Cyclooxygenase-1 or -2--which one mediates lipopolysaccharide-induced hypothermia? Am J Physiol Regul Integr Comp Physio1297: R485--R494, 2009. First published June 10, 2009; doi: 10.1152/ajpregu.91026.2008.--Systemic inflammation is associated with either fever or hypothermia. Fever, a response to mild systemic inflammation, is mediated by cyclooxygenase (COX)-2 and not by COX-1. However, it is still disputed whether COX-2, COX-l, neither, or both mediate(s) responses to severe systemic inflammation, and, in particular, the hypothermic response. We compared the effects of SC-236 (COX-2 inhibitor) and SC-560 (COX-1 inhibitor) on the deep body temperature ([T.sub.b]) of rats injected with a lower (10 [micro]g/kg iv) or higher (1,000 [micro]g/kg iv) dose of LPS at different ambient temperatures ([T.sub.a]s). At a neutral [T.sub.a] (30[degrees]C), the rats responded to LPS with a polyphasic fever (lower dose) or a brief hypothermia followed by fever (higher dose). SC-236 (2.5 mg/kg iv) blocked the fever induced by either LPS dose, whereas SC-560 (5 mg/kg iv) altered neither the febrile response to the lower LPS dose nor the fever component of the response to the higher dose. However, SC-560 blocked the initial hypothermia caused by the higher LPS dose. At a subneutral [T.sub.a] (22[degrees]C), the rats responded to LPS with early (70-90 min, nadir) dose-dependent hypothermia. The hypothermic response to either dose was enhanced by SC-236 but blocked by SC-560. The hypothermic response to the higher LPS dose was associated with a fall in arterial blood pressure. This hypotensive response was attenuated by either SC-236 or SC-560. At the onset of LPS-induced hypothermia and hypotension, the functional activity of the COX-1 pathway (COX-1-mediated [PGE.sub.2] synthesis ex vivo) increased in the spleen but not liver, lung, kidney, or brain. The expression of splenic COX-1 was unaffected by LPS. We conclude that COX-1, but not COX-2, mediates LPS hypothermia, and that both COX isoforms are required for LPS hypotension. body temperature; thermoregulation; fever; inflammation

PG[E.sub.2] produced in the periphery triggers the early phase of the febrile response to infection and may contribute to later phases. It can be hypothesized that peripherally synthesized PG[E.sub.2] transmits febrigenic signals to the brain via vagal afferent nerves. Before testing this hypothesis, we investigated whether the febrigenic effect of intravenously administered PG[E.sub.2] is mediated by the brain and is not the result of a direct action of PG[E.sub.2] on thermoeffectors. In anesthetized rats, intravenously injected PG[E.sub.2] (100 [micro]g/kg) caused an increase in sympathetic discharge to interscapular brown adipose tissue (iBAT), as well as increases in iBAT thermogenesis, end-expired C[O.sub.2], and colonic temperature ([T.sub.c]). All these effects were prevented by inhibition of neuronal function in the raphe region of the medulla oblongata using an intra-raphe microinjection of muscimol. We then asked whether the brain-mediated PG[E.sub.2] fever requires vagal signaling and answered this question by conducting two independent studies in rats. In a study in anesthetized rats, acute bilateral cervical vagotomy did not affect the effects of intravenously injected PG[E.sub.2] (100 [micro]g/kg) on iBAT sympathetic discharge and [T.sub.c]. In a study in conscious rats, administration of PG[E.sub.2] (280 [micro]/kg) via an indwelling jugular catheter caused tail skin vasoconstriction, tended to increase oxygen consumption, and increased [T.sub.c]; none of these responses was affected by total truncal subdiaphragmatic vagotomy performed 2 wk before the experiment. We conclude that the febrile response to circulating PG[E.sub.2] is mediated by the brain, but that it does not require vagal afferent signaling. temperature; thermoregnlation; febrile; vagotomy; raphe; brown adipose tissue

While summarizing the current understanding of how body temperature ([T.sub.b]) is regulated, this review discusses the recent progress in the following areas: central and peripheral thermosensitivity and temperature-activated transient receptor potential (TRP) channels; afferent neuronal pathways from peripheral thermosensors; and efferent thermoeffector pathways. It is proposed that activation of temperature-sensitive TRP channels is a mechanism of peripheral thermosensitivity. Special attention is paid to the functional architecture of the thermoregulatory system. The notion that deep [T.sub.b] is regulated by a unified system with a single controller is rejected. It is proposed that [T.sub.b] is regulated by independent thermoeffector loops, each having its own afferent and efferent branches. The activity of each thermoeffector is triggered by a unique combination of shell and core [T.sub.b]. Temperature-dependent phase transitions in thermosensory neurons cause sequential activation of all neurons of the corresponding thermoeffector loop and eventually a thermoeffector response. No computation of an integrated [T.sub.b] or its comparison with an obvious or hidden set point of a unified system is necessary. Coordination between thermoeffectors is achieved through their common controlled variable, [T.sub.b]. The described model incorporates Kobayashi's views, but Kobayashi's proposal to eliminate the term sensor is rejected. A case against the term set point is also made. Because this term is historically associated with a unified control system, it is more misleading than informative. The term balance point is proposed to designate the regulated level of [T.sub.b] and to attract attention to the multiple feedback, feedforward, and open-loop components that contribute to thermal balance.

Previous studies on the role of cyclooxygenase (COX)-1 and -2 in fever induced by intravenous LPS have failed to investigate the role of these isoenzymes in the earliest responses: monophasic fever (response to a low, near-threshold dose of LPS) and the first phase of polyphasic fever (response to higher doses). We studied these responses in 96 mice that were COX-1 or COX-2 deficient (-/-) or sufficient (+/+). Each mouse was implanted with a temperature telemetry probe into the peritoneal cavity and a jugular catheter. The study was conducted at a tightly controlled, neutral ambient temperature (31[degrees]C). To avoid stress hyperthermia (which masks the onset of fever), all injections were performed through a catheter extension. The +/+ mice responded to intravenous saline with no change in deep body temperature. To a low dose of LPS (1 [micro]g/kg iv), they responded with a monophasic fever. To a higher dose (56 [micro]g/kg), they responded with a polyphasic fever. Neither monophasic fever nor the first phase of polyphasic fever was attenuated in the COX-1 -/- mice, but both responses were absent in the COX-2 -/- mice. The second and third phases of polyphasic fever were also missing in the COX-2 -/- mice. The present study identifies a new, critical role for COX-2 in the mediation of the earliest responses to intravenous LPS: monophasic fever and the first phase of polyphasic fever. It also suggests that no product of the COX-1 gene, including the splice variant COX-1b (COX-3), is essential for these responses. prostaglandin [E.sub.2]; cyclooxygenase-1b; cyclooxygenase- 1[V.sub.1]; cyclooxygenase-3; prostaglandin [H.sub.2] synthase; body temperature; thermoregulation; febrile phases

Most published studies of thermoregulatory responses of mice to LPS involved a stressful injection of LPS, were run at a poorly controlled and often subneutral ambient temperature ([T.sub.a]), and paid little attention to the dependence of the response on the LPS dose. These pitfalls have been overcome in the present study. Male C57BL/6 mice implanted with jugular vein catheters were kept in an environmental chamber at a tightly controlled [T.sub.a]. The relationship between the [T.sub.a]s used and the thermoneutral zone of the mice was verified by measuring tail skin temperature, either by infrared thermography or thermocouple thermometry. Escherichia coli LPS in a wide dose range ([10.sup.0]-[10.sup.4] [micro]g/kg) was administered through an extension of the jugular catheter from outside the chamber. The responses observed were dose dependent. At a neutral [T.sub.a], low (just suprathreshold) doses of LPS ([10.sup.0]-[10.sup.1] [micro]g/kg) caused a monophasic fever. To a slightly higher dose ([10.sup.1.5] [micro]g/kg), the mice responded with a biphasic fever. To even higher doses ([10.sup.1.75]-[10.sup.4] [micro]g/kg), they responded with a polyphasic fever, of which three distinct phases were identified. The dose dependence and dynamics of LPS fever in the mouse appeared to be remarkably similar to those seen in the rat. However, the thermoregulatory response of mice to LPS in a subthermoneutral environment is remarkably different from that of rats. Although very high doses of LPS ([10.sup.4] [micro]g/kg) did cause a late (latency, ~3 h) hypothermic response in mice, the typical early (latency, 10-30 min) hypothermic response seen in rats did not occur. The present investigation identifies experimental conditions to study LPS-induced mono-, bi-, and polyphasic fevers and late hypothermia in mice and provides detailed characteristics of these responses. body temperature; fever; hypothermia; febrile phases; thermoneutrality; systemic inflammation; stress; mice; rats

LPS preparations cause a variety of body temperature ([T.sub.b]) responses: monophasic fever, different phases of polyphasic fever, and hypothermia. Conventional (c) LPS preparations contain highly active lipoprotein contaminants (endotoxin proteins). Whereas LPS signals predominantly via the Toll-like receptor (TLR) 4, endotoxin proteins signal via TLR2. Several TLR2-dependent responses of immunocytes to cLPS in vitro are triggered by endotoxin proteins and not by LPS itself. We tested whether any [T.sub.b] response to cLPS from Escherichia coli 055:B5 is triggered by non-TLR4-signaling contaminants. A decontaminated (d) LPS preparation (free of endotoxin proteins) was produced by subjecting cLPS to phenol-water reextraction. The presence of nonTLR4-signaling contaminants in cLPS (and their absence in dLPS) was confirmed by showing that cLPS (but not dLPS) induced IL-1[beta] expression in the spleen and increased serum levels of TNF-[alpha] and IL-1[beta] of C3H/HeJ mice; these mice bear a nonfunctional TLR4. Yet, both cLPS and dLPS caused cytokine responses in C3H/HeOuJ mice; these mice bear a fully functional TLR4. We then studied the [T.sub.b] responses to cLPS and dLPS in Wistar rats preimplanted with jugular catheters. At a neutral ambient temperature (30[degrees]C), a low (0.1 [micro]g/kg iv) dose of cLPS caused a monophasic fever, whereas a moderate (10 [micro]g/kg iv) dose produced a polyphasic fever. In the cold (20[degrees]C), a high (500 [micro]g/kg iv) dose of cLPS caused hypothermia. All [T.sub.b] responses to dLPS were identical to those of cLPS. We conclude that all known [T.sub.b] responses to LPS preparations are triggered by LPS per se and not by non-TLR4-signaling contaminants of such preparations. body temperature; fever; hypothermia; inflammation; Toll-like receptors; TLR2; TLR4; LPS

In view of the potential involvement of peripherally synthesized, circulating amphipathic mediators [such as platelet-activating factor (PAF) and prostaglandin [E.sub.2]] in the systemic inflammatory response to lipopolysaccharide (LPS), we hypothesized that transport of amphipaths by albumin is essential for conveying peripheral inflammatory signals to the brain. Our first specific aim was to test this hypothesis by studying LPS-induced fever and hypothermia in Nagase analbuminemic rats (NAR). NAR from two different colonies and normalbuminemic Sprague-Dawley rats were preimplanted with jugular catheters, and their febrile responses to a mild dose of LPS (10 [micro]g/kg iv) at thermoneutrality and hypothermic responses to a high dose of LPS (500 [micro]g/kg iv) in the cold were studied. NAR of both colonies developed normal febrile and hypothermic responses, thus suggesting that transport of amphipathic mediators by albumin is not indispensable for LPS signaling. Although alternative carrier proteins [such as [[alpha].sub.1]-acid glycoprotein (AGP)] are known to assume transport functions of albumin in NAR, it is unknown whether inflammatory mediators are capable of inducing their actions when bound to alternative carriers. To test whether PAF, the most potent amphipathic pyrogen, causes fever when administered in an AGP-bound form was our second aim. Sprague-Dawley rats were preimplanted with jugular catheters, and their thermal responses to infusion of a 1:1 [PAF-AGP] complex (40 nmol/kg iv), AGP (40 nmol/kg iv), or various doses of free (aggregated) PAF were studied. The complex, but neither free PAF nor AGP, caused a high (~1.5[degrees]C) fever with a short ( systemic inflammation; fever; hypothermia; body temperature; platelet-activating factor; Nagase analbuminemic rats

Do fever and anapyrexia exist? Analysis of set point-based definitions. Am J Physiol Regul Integr Comp Physiol 287: R992-R995, 2004. First published June 24, 2004; 10.1152/ ajpregu.00068.2004.--Fever and anapyrexia are the most studied thermoregulatory responses. They are defined as a body temperature ([T.sub.b]) increase and decrease, respectively, occurring because of a shift in the set point (SP) and characterized by active defense of the new [T.sub.b]. Although models of [T.sub.b] control with a single SP (whether obvious or hidden) have been criticized, the SP-based definitions have remained unchallenged. In this article, the SP-based definitions of fever and anapyrexia were subjected to two tests. In test 1, they were compared with experimental data on changes in thresholds for activation of different thermoeffectors. Changes in thresholds were found compatible with an SP increase in some (but not all) cases of fever. In all cases of what is called anapyrexia, its mechanism (dissociation of thresholds of different effectors) was found incompatible with a decrease in a single SP. In test 2, experimental data on the dependence of [T.sub.b] on ambient temperature ([T.sub.a]) were analyzed. It was found that the febrile level of [T.sub.b] is defended in some (but not all) cases. However, strong dependence on [T.sub.a] was found in all cases of anapyrexia, which agrees with threshold dissociation but not with a decrease of the SP. It is concluded that fever (as defined) has only limited experimental support, whereas anapyrexia (as defined) does not exist. Two solutions are offered. A palliative is to accept that SP-based terms (anapyrexia, cryexia, regulated hypothermia, and such) are inadequate and should be abandoned. A radical solution is to transform all definitions based on comparing [T.sub.b] with the SP into definitions based on balancing active and passive processes of [T.sub.b] control. thermoregulation; body temperature; febrile response; hypothermia; poikilothermy; thresholds; thermoeffectors; balance point

The organum vasculosum laminae terminalis (OVLT) has been proposed to serve as the interface for blood-to-brain febrigenic signaling, because ablation of this structure affects the febrile response. However, lesioning the OVLT causes many 'side effects' not fully accounted for in the fever literature. By placing OVLT-lesioned rats on intensive rehydration therapy, we attempted to prevent these side effects and to evaluate the febrile response in their absence. After the OVLT of Sprague-Dawley rats was lesioned electrolytically, the rats were given access to 5% sucrose for 1 wk to stimulate drinking. Sucrose consumption and body mass were monitored. The animals were examined twice a day for signs of dehydration and treated with isotonic saline (50 ml/kg sc) when indicated. This protocol eliminated mortality but not several acute and chronic side effects stemming from the lesion. The acute effects included adipsia and gross (14% of body weight) emaciation; chronic effects included hypernatremia, hyperosmolality, a suppressed drinking response to hypertonic saline, and previously unrecognized marked (by ~2[degrees]C) and long-lasting (>3 wk) hyperthermia. Because the hyperthermia was not accompanied by tail skin vasoconstriction, it likely reflected increased thermogenesis. After the rats recovered from the acute (but not chronic) side effects, their febrile response to IL-1[beta] (500 ng/kg iv) was tested. The sham-operated rats developed typical monophasic fevers (~0.5[degrees]C), the lesioned rats did not. However, the absence of the febrile response in the OVLT-lesioned rats likely resulted from the untreatable side effects. For example, hyperthermia at the time of pyrogen injection was high enough (39-40[degrees]C) to solely prevent fever from developing. Hence, the changed febrile responsiveness of OVLT-lesioned animals is given an alternative interpretation, unrelated to febrigenic signaling to the brain. fever; adipsia; third ventricle; median preoptic nucleus

Prostaglandin (PG) [E.sub.2] is a principal downstream mediator of fever and other symptoms of systemic inflammation. Its inactivation occurs in peripheral tissues, primarily the lungs and liver, via carrier-mediated cellular uptake and enzymatic oxidation. We hypothesized that inactivation of PG[E.sub.2] is suppressed during LPS fever and that transcriptional downregulation of PG[E.sub.2] carriers and catabolizing enzymes contributes to this suppression. Fever was induced in inbred Wistar-Kyoto rats by intravenous LPS (50 [micro]g/kg); the controls received saline. Samples of the liver, lungs, and hypothalamus were harvested 0, 0.5, 1.5, and 5 h postinjection. The expression of the two principal transmembrane PG[E.sub.2] carriers (PG transporter and multispecific organic anion transporter) and the two key PG[E.sub.2]-inactivating enzymes [15-hydroxy-PG dehydrogenase (15-PGDH) and carbonyl reductase] was quantified by RT-PCR. All four genes of interest were downregulated in peripheral tissues (but not the brain) during fever. Most remarkably, the expression of hepatic 15-PGDH was decreased 26-fold 5 h post-LPS, whereas expression of pulmonary 15-PGDH was downregulated (as much as 18-fold) throughout the entire febrile course. The transcriptional downregulation of several proteins involved in PG[E.sub.2] inactivation, first reported here, is an unrecognized mechanism of systemic inflammation. By increasing the blood-brain gradient of PG[E.sub.2], this mechanism likely facilitates penetration PG[E.sub.2] into the brain and prevents its elimination from the brain. systemic inflammation; multispecific organic anion transporter; prostaglandin transporter; 15-hydroxyprostaglandin dehydrogenase; carbonyl reductase

The febrile response to lipopolysaccharide (LPS) consists of three phases (phases I-III), all requiring de novo synthesis of prostaglandin (PG) [E.sub.2]. The major mechanism for activation of PG[E.sub.2]-synthesizing enzymes is transcriptional upregulation. The triphasic febrile response of Wistar-Kyoto rats to intravenous LPS (50 [micro]g/kg) was studied. Using real-time RT-PCR, the expression of seven PG[E.sub.2]-synthesizing enzymes in the LPS-processing organs (liver and lungs) and the brain 'febrigenic center' (hypothalamus) was quantified. Phase I involved transcriptional upregulation of the functionally coupled cyclooxygenase (COX)-2 and microsomal (m) PGE synthase (PGES) in the liver and lungs. Phase H entailed robust upregulation of all enzymes of the major inflammatory pathway, i.e., secretory (s) phospholipase (PL) [A.sub.2]-IIA -> COX-2 -> mPGES, in both the periphery and brain. Phase III was accompanied by the induction of cytosolic (c) PL[A.sub.2-[alpha]] in the hypothalamus, further upregulation of sPL[A.sub.2]-IIA and mPGES in the hypothalamus and liver, and a decrease in the expression of COX-1 and COX-2 in all tissues studied. Neither sPL[A.sub.2]-V nor cPGES was induced by LPS. The high magnitude of upregulation of mPGES and sPL[A.sub.2]-IIA (1,257-fold and 133-fold, respectively) makes these enzymes attractive targets for anti-inflammatory therapy. cyclooxygenases; phospholipases; terminal prostaglandin E synthases; lipopolysaccharide; febrile phases

Fever responses of Zucker rats with and without fatty mutation of the leptin receptor. Am J Physiol Regulatory Integrative Comp Physiol 282: R311-R316, 2002; 10.1152/ajpregu. 00376.2001.--Leptin is thought to be involved in febrigenic signaling from the periphery to the brain. Zucker obese rats have a so-called fatty mutation in the leptin receptor gene and express a dysfunctional protein. Studies comparing the fever responses of fatty (fa/fa) rats and of their lean (Fa/Fa and Fa/fa) counterparts yield contradictory results. To resolve these contradictions, we evaluated the effect of fatty mutation on infectious and stress-associated fevers at thermoneutrality (29 [degrees] C) and in a cool environment (20 [degrees] C). Zucker fa/fa and Fa/? rats were infused with Escherichia coli lipopolysaccharide (LPS; 10 [micro]g/kg) through a jugular catheter (infectious fever) or with saline through the catheter (control) or received a painful intramuscular injection of saline (stress fever). At thermoneutrality, the colonic temperature ([T.sub.c]) responses of fatty rats to all stimuli tested were no different from the responses of lean rats. In a cool environment, [T.sub.c] responses of fatty rats to all stimuli were -0.5 [degrees] C lower than those of lean rats. The observed attenuation of LPS-induced and stress-associated fevers in Zucker fatty rats in the cold agrees with the literature data showing that brown adipose tissue (the major heat production effector) is morphologically and functionally defective in these rats. The normal febrile responses of fatty Zucker rats to pyrogenic stimuli at thermoneutrality indicate that fatty mutation does not interrupt febrigenic signaling from the periphery to the brain. infectious fever; stress fever; lipopolysaccharide; endotoxin; thermogenesis; thermoneutrality; cold exposure; obesity

Human serum albumin (HSA) was preincubated into prostaglandin E2 (PGE2) to allow for the formation of a PGE2-HSA complex. The study demonstrated a marked pyrogenic activity of the intravenous PGE2-HSA, but not for the free PGE2. Administration of the free ligand was less physiologically relevant than the administration of a preformed complex because of the ligand's disaggregation, protection from enzymatic degradation and facilitated delivery to targets.

The temperature responses of various rat strains to different lipopolysaccharide (LPS) preparations were monitored to investigate febrile response patterns. Results reveal that instead of the biphasic febrile response in the literature, the subjects exhibited a triphasic response cycle which was independent of the rat strain used and the LPS preparation administered.

Contributory factors that led to the belief that febrile responses are biphasic are examined using the lipopolysaccharide-induced febrile response in rats. Two experiments were conducted to isolate this contributing mechanisms. Results reveal that the initial rise in temperature observed in the triphasic response cannot be differentiated from the temperature rise brought about by stress hyperthermia observed in the control group.

The branch of the vagus nerve responsible for transducing the peripheral signal that induces the elevation of the body temperature during the febrile response to systemic inflammation is investigated by performing selective vagotomies in rats and monitoring their response to low doses of lipopolysaccharides from Escherichia coli. Results reveal that the hepatic branch of the vagus nerve mediates the transduction of peripheral signals in fever genesis.

The cold defense mechanisms in vagotomized rats was examined to test the thermal responsiveness of well-nourished vagotomized rats to various nonpyrogenic stimuli. Results of the study revealed that well-nourished vagotomized rats showed no signs of thermoeffector deficiency. It was concluded that vagal deafferentation is responsible for the febrile irresponsiveness of vagotomized rats.

The febrile responsiveness of vagotomized rats was analyzed to determine whether the vagotomy-induced depression of the febrile responsiveness to pyrogens is a result of missing afferent signals. Results of the investigation indicated that the vagotomy-associated malnutrition can be prevented through perioperative care. Malnutrition is not exactly the reason of the febrile irresponsiveness of vagotomized rats.

Researchers have investigated whether biphasic fever and hypothermia, which are both thermal responses requiring the intact vagus nerve for their development, are vagus mediated and could be affected by subdiaphragmatic vagotomy. This involved assessing thermal responses to the intravenous injection of Escherichia coli in adult Wistar rats 28 days after vagotomy. It was concluded that signals conveyed by the vagus nerve play a key part in initiating monophasic fever.

The pathogenesis and stages of the sickness syndrome during inflammation were investigated in the rat. Analysis of biphasic febrile response of rats to intravenous administration of a bacterial lipopolysaccharide showed that the sickness undergoes two consecutive stages. These stages are critical to the development of systemic inflammatory response and have distinct biological significance.

Endotoxins disrupt may of the body's regulatory mechanisms to an extent that may even lead to death. In rats, endotoxin shock is accompanied by an initial stage of hypotension while endotoxin levels are low. As endotoxin concentration in the body increases, the rat suffers an elevation of core body temperature accompanied by increased oxygen consumption. The rats are also observed to seek out cold temperatures as an effort to alleviate hypothermia.

Intrapreoptic norepinephrine may have a physiological function in the onset of febrilysis in guinea pigs, probably due to its thermolytic influence on fever and on generation of metabolic heat. During the initiation of febrilytic phases, the interstital amount of norepinephrine is increased. A febrilytic action is employed when norepinephrine is microdialyzed in the preoptic region of conscious guinea pigs.