In vivo monitoring of rat brain endocannabinoids using solid-phase microextraction.

Aim: Solid-phase microextraction is proposed to measure concentrations of anandamide and 2-arachidonoyl glycerol in live rat brains in response to stress. Materials & methods: Solid-phase microextraction fibers were prepared from steel with 1.5 mm extraction coating. 24 male rats were divided into groups based on brain region, stria terminalis or posterior hypothalamus and loud noise or control groups. The fibers were desorbed in acetonitrile-water (75:25) and analyzed by ultraperformance LC-MS/MS. The linear range of the method was 0.05-50 ng/ml and the in vivo concentrations were found to be between 0.3 and 40 ng/ml. Conclusion: The new approach was successfully used to determine the concentrations of anandamide and 2-arachidonoyl glycerol in vivo and could be used in the future to measure other endogenous compounds.

The initial fall in arterial pressure evoked by endotoxin is mediated by the ventrolateral periaqueductal gray.

This study tested the hypothesis that the initial fall in arterial pressure evoked by lipopolysaccharide (LPS) is mediated by the ventrolateral column of the midbrain periaqueductal gray region (vlPAG). To test this hypothesis, the local anaesthetic lidocaine (2%; 0.1 µL, 0.2 µL or 1.0 µL), the delta opioid receptor antagonist naltrindole (2 nmol) or saline was microinjected into the vlPAG of isoflurane-anaesthetized rats bilaterally and LPS (1 mg/kg) or saline was administered intravenously 2 min later. Both lidocaine and naltrindole inhibited LPS-evoked hypotension significantly but did not affect arterial pressure in saline-treated control animals. Neither lidocaine nor naltrindole altered heart rate significantly in either LPS-treated or control animals. Microinjection of lidocaine or naltrindole into the dorsolateral PAG was ineffective. These data indicate that the vlPAG plays an important role in the initiation of endotoxic hypotension and further show that delta opioid receptors in the vlPAG participate in the response.

The intracerebroventricular injection of rimonabant inhibits systemic lipopolysaccharide-induced lung inflammation.

We investigated the role of intracerebroventricular (ICV) injection of rimonabant (500ng), a CB1 antagonist, on lipopolysaccharide ((LPS) 5mg/kg)-induced pulmonary inflammation in rats in an isolated perfused lung model. There were decreases in pulmonary capillary pressure (Ppc) and increases in the ((Wet-Dry)/Dry lung weight)/(Ppc) ratio in the ICV-vehicle/LPS group at 4h. There were decreases in TLR4 pathway markers, such as interleukin receptor-associated kinase-1, IκBα, Raf1 and phospho-SFK (Tyr416) at 30min and at 4h increases in IL-6, vascular cell adhesion molecule-1 and myeloperoxidase in lung homogenate. Intracerebroventricular rimonabant attenuated these LPS-induced responses, indicating that ICV rimonabant modulates LPS-initiated pulmonary inflammation.

The OVLT initiates the fall in arterial pressure evoked by high dose lipopolysaccharide: evidence that dichotomous, dose-related mechanisms mediate endotoxic hypotension.

This study tested the hypothesis that lipopolysaccharide (LPS) lowers arterial pressure through two different mechanisms depending on the dose. Previously, we found that a low hypotensive dose of LPS (1mg/kg) lowers arterial pressure by activating vagus nerve afferents. Here we report that hypotension evoked by high dose LPS (15mg/kg) can be prevented by injecting lidocaine into the OVLT but not by vagotomy or inactivation of the NTS. The hypotension produced by both LPS doses was correlated with elevated extracellular norepinephrine concentrations in the POA and prevented by blocking alpha-adrenergic receptors. Thus, initiation of endotoxic hypotension is dose-related, mechanistically.

Alterations in affective behavior during the time course of alcohol hangover.

Alcohol hangover is a temporary state described as the unpleasant next-day effects after binge-like drinking. Hangover begins when ethanol is absent in plasma and is characterized by physical and psychological symptoms. Affective behavior is impaired during the acute phase of alcohol intoxication; however, no reports indicate if similar effects are observed during withdrawal. The aim of this work was to study the time-extension and possible fluctuations in affective behavior during a hangover episode. Male Swiss mice were injected i.p. either with saline (control group) or with ethanol (3.8g/kg BW) (hangover group). Anxiety, fear-related behavior and despair phenotype were evaluated at a basal point (ZT0) and every 2h up to 20h after blood alcohol levels were close to zero (hangover onset). Also, anhedonia signs and pain perception disabilities were studied. Mice exhibited an increase in anxiety-like behavior during 4h and 14h after hangover onset when evaluated by the elevated-plus maze and open field test respectively (p<0.05). Fear-related behavior was detected in hangover animals by the increase of freezing and decrease of line crossings and rearing frequency during 16h after hangover onset (p<0.001). Depression signs were found in hangover mice during 14h (p<0.05). Hangover mice showed a significant decrease in pain perception when tested by tail immersion test at the beginning of hangover (p<0.05). Our findings demonstrate a time-extension between 14 and 16h for hangover affective impairments. This study shows the long lasting effects of hangover over the phase of ethanol intoxication.

Inhibition of monosodium urate crystal-induced inflammation by scopoletin and underlying mechanisms.

The present study determined the anti-inflammatory activity of scopoletin in gout air pouch model and revealed the underlying mechanisms by in vitro assays. Monosodium urate (MSU) crystal-induced inflammation in mouse air pouch model, an experimental model for acute gout, was used to assess the efficacy of scopoletin. The neutrophil and mononuclear phagocyte numbers and MPO levels were increased significantly six hours after MSU crystal injection into the air pouch, whereas these changes were inhibited substantially upon scopoletin (100 and 200mg/kg, i.p.) treatment. To get insight into the underlying mechanisms, the in vitro studies were performed to investigate the effects of scopoletin on activation of macrophages and resultant production of inflammatory mediators. The secretions of interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), prostaglandin E(2) (PGE(2)) and nitric oxide (NO) were elevated in MSU crystal-stimulated RAW 264.7 cells, and scopoletin (30-300 µM) suppressed the production of all mediators. Moreover, RT-PCR assay and western blot analysis indicated that scopoletin regulated the transcriptional level of these mediators via suppression of NF-κB activation and blockade of MAPK signal pathway. Thus, the results clearly indicated that scopoletin inhibited the monosodium urate crystal-induced inflammation both in vivo and in vitro. In combination with our previous findings that scopoletin shows hypouricemic, anti-angiogenesis and pro-apoptotic activities, this compound may be a potential agent for gout therapy and could serve as a structural base for developing new drugs.

Puerarin exerts antipyretic effect on lipopolysaccharide-induced fever in rats involving inhibition of pyrogen production from macrophages.

ETHNOPHARMACOLOGICAL RELEVANCE: Puerarin is the most abundant isoflavonoid in Radix Puerariae (Gegen), which has been prescribed as a medicinal herb for treating fever in China for a long history. AIM OF THE STUDY: The present study aimed at evaluating the antipyretic effect of puerarin and revealing the related mechanisms. MATERIALS AND METHODS: Lipopolysaccharide (LPS)-induced fever in rats was used to assess the antipyretic effect of puerarin. After an intraperitoneal injection of LPS (100µg/kg), body temperature was tested every 30min up to 8h. Different doses of puerarin (25, 50, 100mg/kg) were intraperitoneally administered 30min before LPS injection. In vitro, LPS-stimulated RAW 264.7 cells were treated with various concentrations of puerarin (25-200µM). The pyrogenic mediators, including interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), prostaglandin E(2) (PGE(2)) and nitric oxide (NO), were examined on both transcription and expression levels. Furthermore, the influences of the activation of nuclear factor-kappa B (NF-κB) and the phosphorylation of mitogen-activated protein kinases (MAPKs) by puerarin were assayed by western blot. RESULTS: The intraperitoneal administration of puerarin at test doses clearly demonstrated apparent antipyretic effect through the declines in body temperature elevated by LPS in rats. The in vitro data showed that puerarin inhibited the production of IL-1ß, TNF-α, IL-6, PGE(2) and NO; moreover, the RT-PCR analysis and the western blot analysis indicated that puerarin regulated the transcriptional level via suppression of NF-κB activation and blockade of MAPK signal pathway. CONCLUSIONS: In summary, the antipyretic property of puerarin might result, at least in part, from an inhibition of endogenous pyrogen production and expression. Taken in this sense, our findings provide an explanation for puerarin acting as an important constituent in Gegen, thus, provide scientific basis for the wide use of Radix Puerariae in China as a traditional antipyretic.

Tumor necrosis factor-α induces increased lung vascular permeability: a role for GSK3α/ß.

We tested the hypothesis that glycogen synthase kinase 3α/ß (GSK3α/ß) modulates tumor necrosis factor-a (TNF) induced increased lung vascular permeability. Rats were treated with TNF (i.v., ~100ng/ml) or vehicle 0.5h, 4.0h and 24.0h prior to lung isolation. Rats were co-treated with the GSK3α/ß inhibitors SB216763 (0.6mg/kg) or TDZD-8 (1.0mg/kg). After TNF, the isolated lung was assessed for hemodynamics, wet-dry/dry weight (W-D/D) and extravascular albumin. Extravascular albumin significantly increased at TNF-24h compared to Control. In the GSK3α/ß-inhibited+TNF groups, extravascular albumin was similar to the Control and respective SB216763 and TDZD-8 groups. In separate studies, to assess GSK3α/ß-activity, lung lysate was assessed for phospho-GSK3α/ß-Ser(21/9), total GSK3α/ß, un-phospho-ß-catenin-Ser(33/37) and total ß-catenin. In the TNF-4.0h group, there was no change in GSK3α/phospho-GSK3α-Ser(21) but there was an increase in GSK3ß/GSK3ß-Ser(9) compared to Control, indicating GSK3ß activation at TNF-4.0h. GSK3ß activation was verified because there was a decrease in un-phospho-ß-catenin-Ser(33/37)/ß-catenin in the TNF-4.0 group, a specific outcome for GSK3ß activation. In the SB216763+TNF group, un-phospho-ß-catenin-Ser(33/37) was similar to Control, indicating prevention of TNF-induced GSK3ß activation. In the TNF-24h group, there were increases in the biomarkers of inflammation phospho-eNOS-Ser (1117) and oxidized protein, which did not occur in the SB216763+TNF-24h and TDZD-8+TNF-24h groups. In the SB216763+TNF-24h and TDZD-8+TNF-24h groups, un-phospho-ß-catenin-Ser(33/37) was greater than in the Control, indicating continued inhibition of GSK3ß. The data indicates that pharmacologic inhibition of GSK3ß inhibits TNF induced increased endothelial permeability associated with lung inflammation.

Food deprivation alters thermoregulatory responses to lipopolysaccharide by enhancing cryogenic inflammatory signaling via prostaglandin D2.

We tested the hypothesis that food deprivation alters body temperature (T(b)) responses to bacterial LPS by enhancing inflammatory signaling that decreases T(b) (cryogenic signaling) rather than by suppressing inflammatory signaling that increases T(b) (febrigenic signaling). Free-feeding or food-deprived (24 h) rats received LPS at doses (500 and 2,500 microg/kg iv) that are high enough to activate both febrigenic and cryogenic signaling. At these doses, LPS caused fever in rats at an ambient temperature of 30 degrees C, but produced hypothermia at an ambient temperature of 22 degrees C. Whereas food deprivation had little effect on LPS fever, it enhanced LPS hypothermia, an effect that was particularly pronounced in rats injected with the higher LPS dose. Enhancement of hypothermia was not due to thermogenic incapacity, since food-deprived rats were fully capable of raising T(b) in response to the thermogenic drug CL316,243 (1 mg/kg iv). Neither was enhancement of hypothermia associated with altered plasma levels of cytokines (TNF-alpha, IL-1beta, and IL-6) or with reduced levels of an anti-inflammatory hormone (corticosterone). The levels of PGD(2) and PGE(2) during LPS hypothermia were augmented by food deprivation, although the ratio between them remained unchanged. Food deprivation, however, selectively enhanced the responsiveness of rats to the cryogenic action of PGD(2) (100 ng icv) without altering the responsiveness to febrigenic PGE(2) (100 ng icv). These findings support our hypothesis and indicate that cryogenic signaling via PGD(2) underlies enhancement of LPS hypothermia by food deprivation.

Neonatal intrahippocampal immune challenge alters dopamine modulation of prefrontal cortical interneurons in adult rats.

BACKGROUND: Among the diverse animal models proposed for schizophrenia, the neonatal ventral hippocampal lesion (NVHL) is one of the most widely used. However, its construct validity can be questioned because there is no evidence of a lesion present in schizophrenia. Other approaches that have tried to capture environmental influences on development include diverse models of maternal infection. METHODS: As the early postnatal days in rodents are equivalent to the third trimester of human pregnancy in terms of brain development, we decided to test whether a neonatal immune challenge with an injection of the bacterial endotoxin lipopolysaccharide (LPS) into the ventral hippocampus caused deficits in interneuron function similar to those reported for the NVHL. RESULTS: Neonatal LPS injection caused a persistent elevation in cytokines in several brain regions, deficits in prepulse inhibition of the acoustic startle response, and a loss of the periadolescent maturation in the response of prefrontal cortical fast-spiking interneurons to dopamine. CONCLUSIONS: The same phenotypes elicited by a NVHL can be obtained with an intrahippocampal immune challenge, suggesting that perinatal environmental factors can affect adult prefrontal interneuron maturation during adolescence.

Central cannabinoid 1 receptor antagonist administration prevents endotoxic hypotension affecting norepinephrine release in the preoptic anterior hypothalamic area.

It is widely assumed that LPS lowers arterial pressure during sepsis by stimulating release of TNF-alpha and other vasoactive mediators from macrophages. However, recent data from this and other laboratories have shown that LPS hypotension can be prevented by inhibiting afferent impulse flow in the vagus nerve, by blocking neuronal activity in the nucleus of the solitary tract, or by blocking alpha-adrenergic receptors in the preoptic area/anterior hypothalamic area (POA). These findings suggest that the inflammatory signal is conveyed from the periphery to the brain via the vagus nerve, and that endotoxic shock is mediated through a central mechanism that requires activation of POA neurons. In the present study, we tested whether central cannabinoid 1 (CB1) receptors participate in the control of arterial pressure during endotoxemia based on evidence that hypothalamic neurons express CB1 receptors and synthesize the endogenous CB anandamide. We found that intracerebroventricular administration of rimonabant, a CB1 receptor antagonist, inhibited the fall in arterial pressure evoked by LPS significantly in both conscious and anesthetized rats. Rimonabant attenuated both the immediate fall in arterial pressure evoked by LPS and the second, delayed hypotensive phase that leads to tissue ischemia and death. Rimonabant also prevented the associated LPS-induced rise in extracellular fluid norepinephrine concentrations in the POA. Furthermore, rimonabant attenuated the associated increase in plasma TNF-alpha concentrations characteristic of the late phase of endotoxic hypotension. These data indicate that central CB1 receptors may play an important role in the initiation of endotoxic hypotension.

Lipopolysaccharide-induced hypotension is mediated by a neural pathway involving the vagus nerve, the nucleus tractus solitarius and alpha-adrenergic receptors in the preoptic anterior hypothalamic area.

We recently reported that the preoptic anterior hypothalamic area (POA) mediates the hypotensive response evoked by lipopolysaccharide (LPS). In this study, we investigated how the inflammatory signal induced by LPS reaches the POA. Subdiaphragmatic vagotomy and abdominal perivagal lidocaine administration, or lidocaine injection into the nucleus tractus solitarius (NTS) prevented LPS hypotension. Microinjection of the alpha-adrenergic receptor antagonist phentolamine into the POA, blocked initiation of the hypotensive response and prevented the late decompensatory phase. These data suggest that LPS hypotension is mediated by the vagus nerve which conveys the signal to the NTS and, in turn, stimulates norepinephrine release within the POA.

The preoptic anterior hypothalamic area mediates initiation of the hypotensive response induced by LPS in male rats.

The mechanism responsible for the initiation of endotoxic hypotension is not fully understood, although it is often attributed to a direct effect of LPS and other vasoactive mediators on the vasculature. Alternatively, recent evidence raises the possibility that endotoxic hypotension may be initiated through a central mechanism. Previous studies have shown that LPS initiates fever, sickness behavior, and other aspects of the inflammatory response through a neural pathway that sends peripheral inflammatory signals to the preoptic anterior hypothalamic area (POA). It is also well known that the POA plays a role in the regulation of cardiovascular function, but its involvement in LPS-induced hypotension has not been examined previously. Therefore, the aim of the present paper was to investigate whether the initial abrupt fall in arterial pressure evoked by LPS in septic shock is mediated by the POA. LPS (1 mg/kg, i.v.) administration to halothane-anesthetized or conscious rats lowered arterial blood pressure by 24.8+/-2.9 and 25.1+/-5.8 mmHg, respectively. Bilateral lidocaine (2%; 1 microL) injection into the POA, but not the lateral hypothalamus, prevented the hypotension evoked by LPS entirely in both anesthetized and conscious animals. Remarkably, this blockade significantly inhibited the second, delayed fall in arterial pressure induced by LPS, and simultaneously decreased TNF-alpha plasma levels. Together, these data indicate that the initial phase of endotoxic hypotension is mediated by the POA and suggest that the initiation of the hypotensive response induced by LPS can be essential for the development of the late fall in blood pressure.

Effects of prenatal stress on motor performance and anxiety behavior in Swiss mice.

Stressor presence during the last weeks of gestation has been associated with behavioral disorders in later life. In this study we support further research on the long term effects of prenatal stress on Swiss mice descendant's behavior. Prenatal stress procedure consisted on restraining the dams under bright light for 45 min, three times per day from the 15th day of pregnancy, until birth. After weaning, offspring's motor performance and spontaneous exploratory behavior were measured by the tight-rope and T-maze tests, respectively. We also evaluated anxiety behavior using elevated plus maze test. We found that maternal stress improves the performance of the animals in the tight rope test and that this effect was sex and age dependent: prenatal stressed males obtained the best scores during the first month of life, while in females the same was achieved at the second month. Spontaneous exploratory behavior analysis revealed that it was elevated in prenatal stressed males and that this effect persisted on time. However, we did not find significant differences on this behavioral response among both females groups. Finally, differences on anxiety behavior were found only in females: prenatally stressed animals showed a higher proportion of entries into the open arms of a plus maze (reduced anxiety) compared to the control group. Our results show that prenatal stress modifies the normal behavior of the progeny: prenatal stressed animals have a better performance in the carried out test. These notably results suggest the existence of an adaptive response to prenatal stress.

Preoptic nitric oxide attenuates endotoxic fever in guinea pigs by inhibiting the POA release of norepinephrine.

Lipopolysaccharide (LPS) administration induces hypothalamic nitric oxide (NO); NO is antipyretic in the preoptic area (POA), but its mechanism of action is uncertain. LPS also stimulates the release of preoptic norepinephrine (NE), which mediates fever onset. Because NE upregulates NO synthases and NO induces cyclooxygenase (COX)-2-dependent PGE(2), we investigated whether NO mediates the production of this central fever mediator. Conscious guinea pigs with intra-POA microdialysis probes received LPS intravenously (2 mug/kg) and, thereafter, an NO donor (SIN-1) or scavenger (carboxy-PTIO) intra-POA (20 mug/mul each, 2 mul/min, 6 h). Core temperature (T(c)) was monitored constantly; dialysate NE and PGE(2) were analyzed in 30-min collections. To verify the reported involvement of alpha(2)-adrenoceptors (AR) in PGE(2) production, clonidine (alpha(2)-AR agonist, 2 mug/mul) was microdialyzed with and without SIN-1 or carboxy-PTIO. To assess the possible involvement of oxidative NE and/or NO products in the demonstrated initially COX-2-independent POA PGE(2) increase, (+)-catechin (an antioxidant, 3 mug/mul) was microdialyzed, and POA PGE(2), and T(c) were determined. SIN-1 and carboxy-PTIO reduced and enhanced, respectively, the rises in NE, PGE(2), and T(c) produced by intravenous LPS. Similarly, they prevented and increased, respectively, the delayed elevations of PGE(2) and T(c) induced by intra-POA clonidine. (+)-Catechin prevented the LPS-induced elevation of PGE(2), but not of T(c). We conclude that the antipyretic activity of NO derives from its inhibitory modulation of the LPS-induced release of POA NE. These data also implicate free radicals in POA PGE(2) production and raise questions about its role as a central LPS fever mediator.

Preoptic norepinephrine mediates the febrile response of guinea pigs to lipopolysaccharide.

Norepinephrine (NE) microdialyzed in the preoptic area (POA) raises core temperature (T(c)) via 1) alpha(1)-adrenoceptors (AR), quickly and independently of POA PGE(2), and 2) alpha(2)-AR, after a delay and PGE(2) dependently. Since systemic lipopolysaccharide (LPS) activates the central noradrenergic system, we investigated whether preoptic NE mediates LPS fever. We injected LPS (2 microg/kg iv) in guinea pigs prepared with intra-POA microdialysis probes and determined POA cerebrospinal (CSF) NE levels. We similarly microdialyzed prazosin (alpha(1) blocker, 1 microg/microl), yohimbine (alpha(2) blocker, 1 microg/microl), SC-560 [cyclooxygenase (COX)-1 blocker, 5 microg/microl], acetaminophen (presumptive COX-1v blocker, 5 microg/microl), or MK-0663 (COX-2 blocker, 0.5 microg/microl) in other animals before intravenous LPS and measured CSF PGE(2). All of the agents were perfused at 2 microg/min for 6 h. T(c) was monitored constantly. POA NE peaked within 30 min after LPS and then returned to baseline over the next 90 min. T(c) increased within 12 min to a first peak at approximately 60 min and to a second at approximately 150 min and then declined over the following 2.5 h. POA PGE(2) followed a concurrent course. Prazosin pretreatment eliminated the first T(c) rise but not the second; PGE(2) rose normally. Yohimbine pretreatment did not affect the first T(c) rise, which continued unchanged for 6 h; the second rise, however, was absent, and PGE(2) levels did not increase. SC-560 and acetaminophen did not alter the LPS-induced PGE(2) and T(c) rises; MK-0663 prevented both the late PGE(2) and T(c) rises. These results confirm that POA NE is pivotal in the development of LPS fever.

Kupffer cell-generated PGE2 triggers the febrile response of guinea pigs to intravenously injected LPS.

Because the onset of fever induced by intravenously (i.v.) injected bacterial endotoxic lipopolysaccharides (LPS) precedes the appearance in the bloodstream of pyrogenic cytokines, the presumptive peripheral triggers of the febrile response, we have postulated previously that, in their stead, PGE2 could be the peripheral fever trigger because it appears in blood coincidentally with the initial body core temperature (Tc) rise. To test this hypothesis, we injected Salmonella enteritidis LPS (2 microg/kg body wt i.v.) into conscious guinea pigs and measured their plasma levels of LPS, PGE2, TNF-alpha, IL-1beta, and IL-6 before and 15, 30, 60, 90, and 120 min after LPS administration; Tc was monitored continuously. The animals were untreated or Kupffer cell (KC) depleted; the essential involvement of KCs in LPS fever was shown previously. LPS very promptly (<10 min) induced a rise of Tc that was temporally correlated with the elevation of plasma PGE2. KC depletion prevented the Tc and plasma PGE2 rises and slowed the clearance of LPS from the blood. TNF-alpha was not detectable in plasma until 30 min and in IL-1beta and IL-6 until 60 min after LPS injection. KC depletion did not alter the times of appearance or magnitudes of rises of these cytokines, except TNF-alpha, the maximal level of which was increased approximately twofold in the KC-depleted animals. In a follow-up experiment, PGE2 antiserum administered i.v. 10 min before LPS significantly attenuated the febrile response to LPS. Together, these results support the view that, in guinea pigs, PGE2 rather than pyrogenic cytokines is generated by KCs in immediate response to i.v. LPS and triggers the febrile response.

Cytokines, PGE2 and endotoxic fever: a re-assessment.

The innate immune system serves as the first line of host defense against the deleterious effects of invading infectious pathogens. Fever is the hallmark among the defense mechanisms evoked by the entry into the body of such pathogens. The conventional view of the steps that lead to fever production is that they begin with the biosynthesis of pyrogenic cytokines by mononuclear phagocytes stimulated by the pathogens, their release into the circulation and transport to the thermoregulatory center in the preoptic area (POA) of the anterior hypothalamus, and their induction there of cyclooxygenase (COX)-2-dependent prostaglandin (PG)E(2), the putative final mediator of the febrile response. But data accumulated over the past 5 years have gradually challenged this classical concept, due mostly to the temporal incompatibility of the newer findings with this concatenation of events. Thus, the former studies generally overlooked that the production of cytokines and the transduction of their pyrogenic signals into fever-mediating PGE(2) proceed at relatively slow rates, significantly slower certainly than the onset latency of fever produced by the i.v. injection of bacterial endotoxic lipopolysaccharides (LPS). Here, we review the conflicts between the earlier and the more recent findings and summarize new data that reconcile many of the contradictions. A unified model based on these data explicating the generation and maintenance of the febrile response is presented. It postulates that the steps in the production of LPS fever occur in the following sequence: the immediate activation by LPS of the complement (C) cascade, the stimulation by the anaphylatoxic C component C5a of Kupffer cells, their consequent, virtually instantaneous release of PGE(2), its excitation of hepatic vagal afferents, their transmission of the induced signals to the POA via the ventral noradrenergic bundle, and the activation by the thus, locally released norepinephrine (NE) of neural alpha(1)- and glial alpha(2)-adrenoceptors. The activation of the first causes an immediate, PGE(2)-independent rise in core temperature (T(c)) [the early phase of fever; an antioxidant-sensitive PGE(2) rise, however, accompanies this first phase], and of the second a delayed, PGE(2)-dependent T(c) rise [the late phase of fever]. Meanwhile-generated pyrogenic cytokines and their consequent upregulation of blood-brain barrier cells COX-2 also contribute to the latter rise. The consecutive steps that initiate the febrile response to LPS would now appear, therefore, to occur in an order different than conceived originally.

Putative antihyperpyretic factor induced by LPS in spleen of guinea pigs.

We reported previously that the onset of LPS-induced fever, irrespective of its route of administration, is temporally correlated with the appearance of LPS in the liver and that splenectomy significantly increases both the febrile response to LPS and the uptake of LPS by Kupffer cells (KC). To further evaluate the role of the spleen in LPS fever production, we ligated the splenic vein and, 7 and 30 days later, monitored the core temperature changes over 6 h after intraperitoneal (ip) injection of LPS (2 microg/kg). Both the febrile response and the uptake of LPS by KC were significantly augmented. Like splenectomy, splenic vein ligation (SVL) increased the febrile response and LPS uptake by KC until the collateral circulation developed, suggesting that the spleen may normally contribute an inhibitory factor that limits KC uptake of LPS and thus affects the febrile response. Subsequently, to verify the presence of this factor, we prepared splenic extracts from guinea pigs pretreated with LPS (8 microg/kg ip) or pyrogen-free saline, homogenized and ultrafiltered them, and injected them intravenously into splenectomized (Splex) guinea pigs pretreated with LPS (8 microg/kg ip). The results confirmed our presumption that the splenic extract from LPS-treated guinea pigs inhibits the exaggerated febrile response and the LPS uptake by the liver of Splex guinea pigs, indicating the presence of a putative splenic inhibitory factor, confirming the participation of the spleen in LPS-induced fever, and suggesting the existence of a novel antihyperpyretic mechanism. Preliminary data indicate that this factor is a lipid.

Lipopolysaccharide challenge causes exaggerated fever and increased hepatic lipopolysaccharide uptake in vinblastine-induced leukopenic guinea pigs.

OBJECTIVE: To better understand the pathophysiology of the fever often manifested by immunocompromised patients undergoing chemotherapy that become neutropenic and suffer a bacterial infection. DESIGN: Prospective animal study. SETTING: A physiology laboratory in a medical school setting. MEASUREMENTS AND MAIN RESULTS: We induced leukopenia in guinea pigs with vinblastine (0.7 mg/kg, intravenously, 4 days before) and measured the animals' febrile response to 2 microg of lipopolysaccharide/kg and the uptake of 75 microg of fluorescein isothiocyanate-labeled lipopolysaccharide/kg by Kupffer cells. The leukopenic animals exhibited significantly higher fevers and greater hepatic fluorescein isothiocyanate-lipopolysaccharide uptake than their controls. CONCLUSION: Lipopolysaccharide-challenged, vinblastine-induced leukopenic guinea pigs exhibit hyperpyrexia and significantly elevated uptake of lipopolysaccharide by Kupffer cells, the major source of pyrogenic mediators. This could explain "febrile neutropenia."