AIM:Brown adipose tissue thermogenesis is widely considered an important effector mechanism in fever and emotionally induced hyperthermia (psychogenic fever), and increases in interscapular temperature are commonly interpreted as evidence of brown adipose tissue activation. Yet direct evidence for this interpretation remains limited and conflicting. Here, we tested these assumptions using complementary genetic and surgical approaches in rats. METHODS:We generated uncoupling protein 1 knockout rats, in which canonical brown adipose tissue thermogenesis is abolished, and examined body temperature responses to systemic inflammation induced by lipopolysaccharide and to emotional stressors, including restraint and cage exchange. In separate experiments, interscapular brown adipose tissue was surgically removed in wild-type rats. RESULTS:Despite profound impairment of cold- and β₃-adrenergic-induced thermogenesis, uncoupling protein 1 deletion did not affect lipopolysaccharide- or stress-induced increases in core or interscapular temperature. Likewise, removal of interscapular brown adipose tissue did not alter the interscapular temperature response. Thus, increases in interscapular temperature occurred in the absence of both uncoupling protein 1 and interscapular brown adipose tissue. Consistent with these findings, immune challenge and emotional stress induced little or no change in thermogenic gene expression and had minimal effects on tissue mass, in contrast to robust activation by β₃-adrenergic stimulation. CONCLUSION:Fever and psychogenic hyperthermia in rats do not require uncoupling protein 1-mediated brown adipose tissue thermogenesis. Furthermore, increases in interscapular temperature are not necessarily indicative of brown adipose tissue thermogenesis, challenging a widely held interpretation of these phenomena.

Brown adipose tissue (BAT) thermogenesis is essential for cold defense, but its contribution to fever and emotionally induced hyperthermia (psychogenic fever) remains disputed. Here we address this issue using genetic, surgical, physiological, and molecular approaches in rats. We generated UCP1 knockout rats, in which classical BAT thermogenesis is abolished, and examined body temperature responses to systemic inflammation induced by lipopolysaccharide and to emotional stressors such as restraint and cage exchange. Despite profound impairment of cold-induced and β₃-adrenergic-induced thermogenesis, UCP1 deletion did not affect LPS- or stress-evoked elevations in core or interscapular temperature. Surgical removal of interscapular BAT in wild-type rats likewise failed to alter these hyperthermic responses. Consistent with these findings, LPS and emotional stress induced only small, strain-dependent changes in the expression of thermogenic genes in BAT and minimally affected BAT mass, in marked contrast to the robust BAT activation elicited by β₃-adrenergic stimulation. Notably, emotional stress induced UCP3 expression in neck muscles, suggesting a potential contribution of skeletal muscle metabolic processes to stress-induced hyperthermia. Together, these findings demonstrate that both immune-induced and psychogenic fever occur independently of BAT thermogenesis and point to non-BAT tissues - likely including skeletal muscle - as candidate peripheral effectors supporting fever and emotional hyperthermia. ### Competing Interest Statement The authors have declared no competing interest. JSPS KAKENHI, 22H00396, 23KK0126 Swedish Brain Foundation, FO2025-0110-HK-175

Fever induced by systemic inflammation is mediated by prostaglandin E2 (PGE2), but the identity of the prostaglandin E (EP) receptor subtype responsible for the febrile response remains debated. Although EP3 receptors have been strongly implicated in fever generation, studies using pharmacological approaches and receptor-deficient mice have also suggested involvement of EP1 and EP4 receptors. Because previous studies have differed substantially with regard to experimental conditions, including ambient temperature, route of immune challenge, and injection-associated stress responses, we systematically compared the febrile response to lipopolysaccharide (LPS) in mice lacking EP1, EP2, EP3, or EP4 receptors under standardized physiological conditions. Body temperature was recorded by telemetry in mice housed at thermoneutrality and injected intravenously with LPS (30 μg/kg) through indwelling jugular catheters, permitting remote injections with minimal disturbance to the animals. Mice with global deletion of EP1, EP2, or EP3, as well as mice with nervous system-directed deletion of EP4, all maintained on a C57BL/6 background, were examined together with wild-type littermates. Wild-type mice displayed a characteristic multiphasic fever following LPS administration. Mice lacking EP1 or EP2, or with nervous system-directed deletion of EP4, exhibited febrile responses indistinguishable from those of their wild-type controls. In contrast, mice lacking EP3 failed completely to develop fever and instead displayed a pronounced hypothermic response immediately following LPS injection. These findings identify EP3 as the only prostaglandin E receptor with a non-redundant role in LPS-induced fever and support the concept that EP3-dependent signaling constitutes the final common pathway for inflammatory fever.

A.D. (Bud) Craig (1951-2023) redefined the concept of interoception and provided a novel, revolutionary understanding of the neural basis for human awareness. In unsurpassed anatomical-physiological studies in monkeys, Craig showed that the insular cortex is the primary sensory cortex for interoception, or the image of the "material me" that provides a homeostatic representation of the physiological condition of the body. He showed that the insula contains a postero-anteriorly organized somatotopic map of the interoceptive sensations, and that it encodes both the localization and the intensity discrimination of interoceptive sensations. In seminal work in humans, he demonstrated that the interoceptive feelings are re-represented, and multimodally integrated, in anterior portions of the insula in sequence of increasingly homeostatically efficient representations that integrate all salient neural activity. He further showed that subjective awareness is associated with activation of the anterior insular cortex and suggested that this brain region also is critical for fluid intelligence and the perception of time. His work has led to a paradigm shift in our understanding of interoception and how interoceptive sensations underlie consciousness, a topic that long has been considered elusive, or even beyond our comprehension.

Our body temperature is normally kept within a narrow range of 1°C. For example, if our body temperature rises, such as in a hot environment or due to strenuous exercise, our thermoregulatory system will trigger a powerful heat defense response with vasodilation, sweating, and lowered metabolism. During fever, which often involves body temperatures of up to 41°C, this heat defense mechanism is apparently inhibited; otherwise, the rising body temperature would be immediately combated, and fever would not be allowed to develop. New evidence suggests how and where this inhibition takes place. In two consecutive studies from Cheng et al. and Xu et al., it has been shown that prostaglandin E2, which generates fever by acting on thermosensory neurons in the preoptic hypothalamus, also acts on neurons in the brainstem parabrachial nucleus, which receive temperature information from temperature-activated spinal cord neurons and relay this information to the thermoregulatory center in the hypothalamus to either induce cold or heat defenses. By acting on the same type of prostaglandin E2 receptor that is critical for fever generation in the preoptic hypothalamus, the EP3 receptor, prostaglandin E2 inhibits the signaling of the heat-responsive parabrachial neurons, while stimulating the cold-responsive neurons. These novel findings thus show that prostaglandin E2, by binding to the same receptor subtype in the parabrachial nucleus as in the preoptic hypothalamus, adjusts the sensitivity of the thermosensory system in a coordinated manner to allow the development of febrile body temperatures.

The ability to detect and respond to sickness in others promotes survival. Here we show that mouse dams respond to immune challenged pups by mirroring their inflammatory response. Dams with pups subjected to immune challenge displayed a marked induction of inflammatory mediators in both the brain and the periphery, accompanied by an increase in maternal behaviors and corticosterone levels. This social transmission of inflammation did not require physical contact, and it contributed to the stress hormone response in the dams. In adult dyads, interaction with an immune challenged cagemate did not elicit robust inflammatory signaling but induced an increased responsiveness to a subsequent immune challenge. The identification of social transmission of inflammation, or inflammatory responsiveness, may open new avenues for research on social behavior, just like the description of similar phenomena such as observational fear and transmitted pain has done.

Strigo, Irina A.; Andrew, David; Simmons, Alan N.; Evrard, Henry C.; Blomqvist, Anders; Dostrovsky, Jonathan O. Author Information

Letter to the Editor: We concur with Luis Garcia-Larrea9 that the article by Mandonnet et al.12 represents a valuable contribution to our understanding of the role of the insula for our pain perception, most critically because it conclusively shows in humans that the nociceptive input to the dorsal posterior insular cortex is not relayed there through other cortical regions, as was long suggested,13,16 but arrive through ascending fibers from the thalamus. However, this important contribution is, quite unnecessarily, tainted by the authors' failure to name the thalamic nucleus that projects to the posterior insula, writing instead that “it has been suggested that a subpart of the Po-SG specifically projects to an area in the posterior dorsal fundus in the insula,” despite the fact that this “subpart” was identified as a distinct nucleus several decades ago and was designated the VMpo (the posterior part of the ventromedial nucleus).6 It was given this name, which adheres to the commonly used nomenclature for the mammalian thalamus,10 because it lies adjacent (and posterior) to the VMb (the basal part of the ventromedial nucleus), together with which it provides a homeostatic afferent input to the insular cortex, that from VMpo arising from neurons in the spinal and trigeminal superficial dorsal horn and that from VMb from the nucleus of the solitary tract.3 Tract-tracing and electrophysiological recordings in macaque monkeys clearly identified VMpo, and neither the suprageniculate (SG) nor the posterior nucleus (Po), as the target of spinal lamina I projection neurons encoding pain and temperature modalities2,7 and as the source of thalamocortical projections to the dorsal posterior fundus of the insula.4 VMpo is readily identifiable in brain sections both from monkey and human thalamus1,2 (Fig. 1).Figure 1.: Adjacent frontal sections through the mesodiencephalic border zone of the human brain, stained with thionin (A) and for calbindin immunoreactivity (B). Dorsal is upward and lateral is to the left. Arrowheads in (A) indicate the borders of the VMpo nucleus. Note the medial lemniscus (ml) ventral to VMpo and heading toward the ventroposterior nuclei. Panel (B) shows how calbindin-positive fibres, identifying the lamina I spinothalamic tract,7 from the spinal lemniscus (arrowheads in B) enter the VMpo nucleus and form dense terminal-like patches. The same capillaries are indicated by “x” in each pair of images for orientation. Li, nucleus limitans; MG, medial geniculate nucleus; Pla, anterior pulvinar; Sg, suprageniculate nucleus; VMpo, posterior part of the ventromedial nucleus; VPL, ventroposterolateral nucleus. Reprinted from Ref. 1; with permission.We are certainly aware that some 20 years ago, when the available data were still incomplete, influential investigators put a curse on the VMpo by questioning its existence (and consequently also the critical role of its insular projection for the perception of pain)11,16 (but see Ref. 5). However, as pointed out by Vierck et al.15 already in 2013, the arguments that were put forward against VMpo were since long dated, being superseded by subsequent evidence, and became even more so in following years. Today, the role of VMpo as a dedicated thalamoinsular relay for pain and temperature, and other information on the condition of the body, from lamina I of the spinal and trigeminal dorsal horn is thoroughly established.3,4,8,14,15 We therefore believe that it is long past time to cite the seminal studies demonstrating for the first time the existence of the primate spino-thalamo-insular pathway that has become so crucial to the field of pain9,12 and to recognize the VMpo in this context. Conflict of interest statement The authors declare no conflicts of interest.

We recently demonstrated that prostaglandin production in brain endothelial cells is both necessary and sufficient for the generation of fever during systemic immune challenge. I here discuss this finding in light of the previous literature and point to some unresolved issues.

Anorexia is a common symptom during infectious and inflammatory disease. Here we examined the role of melanocortin-4 receptors (MC4Rs) in inflammation-induced anorexia. Mice with transcriptional blockage of the MC4Rs displayed the same reduction of food intake following peripheral injection of lipopolysaccharide as wild type mice but were protected against the anorexic effect of the immune challenge in a test in which fasted animals were to use olfactory cues to find a hidden cookie. By using selective virus-mediated receptor re-expression we demonstrate that the suppression of the food-seeking behavior is subserved by MC4Rs in the brain stem parabrachial nucleus, a central hub for interoceptive information involved in the regulation of food intake. Furthermore, the selective expression of MC4R in the parabrachial nucleus also attenuated the body weight increase that characterizes MC4R KO mice. These data extend on the functions of the MC4Rs and show that MC4Rs in the parabrachial nucleus are critically involved in the anorexic response to peripheral inflammation but also contribute to body weight homeostasis during normal conditions.

Bud Craig, an outstanding neuroscientist, died on 15 July 2023 at age 71. Bud made unique contributions to the fields of pain and interoception, challenging major dogmas and offering powerful explanations for various phenomena including central pain and the subjective awareness of feelings, with great implications for our understanding of consciousness.

ABSTRACT The initiation of fever has been a matter of controversy. Based on observations of little or no induction of prostaglandin synthesizing enzymes in the brain during the first phase of fever it was suggested that fever is initiated by prostaglandin released into the circulation from cells in the liver and lungs. Here we show in the mouse that prostaglandin synthesis is rapidly induced in the brain after immune challenge. These data are consistent with our recent findings in functional experiments that prostaglandin production in brain endothelial cells is both necessary and sufficient for the generation of all phases of fever.

Fever is known to be elicited by prostaglandin E2 acting on the brain, but its origin has remained disputed. We show in mice that selective deletion of prostaglandin synthesis in brain endothelial cells, but not in neural cells or myeloid cells, abolished fever induced by intravenous administration of lipopolysaccharide and that selective rescue of prostaglandin synthesis in brain endothelial cells reinstated fever. These data demonstrate that prostaglandin production in brain endothelial cells is both necessary and sufficient for eliciting fever.

We examined the signaling route for fever during localized inflammation in male and female mice, elicited by casein injection into a preformed air pouch. The localized inflammation gave rise to high concentrations of prostaglandins of the E species (PGE2) and cytokines in the air pouch and elevated levels of these inflammatory mediators in plasma. There were also elevated levels of PGE2 in the cerebrospinal fluid, although there was little evidence for PGE2 synthesis in the brain. Global deletion of the PGE2 prostaglandin E receptor 3 (EP3) abolished the febrile response as did deletion of the EP3 receptor in neural cells, whereas its deletion on peripheral nerves had no effect, implying that PGE2 action on this receptor in the CNS elicited the fever. Global deletion of the interleukin-1 receptor type 1 (IL-1R1) also abolished the febrile response, whereas its deletion on neural cells or peripheral nerves had no effect. However, deletion of the IL-1R1 on brain endothelial cells, as well as deletion of the interleukin-6 receptor α on these cells, attenuated the febrile response. In contrast, deletion of the PGE2 synthesizing enzymes cyclooxygenase-2 and microsomal prostaglandin synthase-1 in brain endothelial cells, known to attenuate fever evoked by systemic inflammation, had no effect. We conclude that fever during localized inflammation is not mediated by neural signaling from the inflamed site, as previously suggested, but is dependent on humoral signaling that involves interleukin actions on brain endothelial cells, probably facilitating PGE2 entry into the brain from the circulation and hence representing a mechanism distinct from that at work during systemic inflammation.

Paracetamol, one of the most widely used pain-relieving drugs, is deacetylated to 4-aminophenol (4-AP) that undergoes fatty acid amide hydrolase (FAAH)-dependent biotransformation into N-arachidonoylphenolamine (AM404), which mediates TRPV1-dependent antinociception in the brain of rodents. However, paracetamol is also converted to the liver-toxic metabolite N-acetyl-p-benzoquinone imine already at therapeutic doses, urging for safer paracetamol analogues. Primary amine analogues with chemical structures similar to paracetamol were evaluated for their propensity to undergo FAAH-dependent N-arachidonoyl conjugation into TRPV1 activators both in vitro and in vivo in rodents. The antinociceptive and antipyretic activity of paracetamol and primary amine analogues was examined with regard to FAAH and TRPV1 as well as if these analogues produced acute liver toxicity. 5-Amino-2-methoxyphenol (2) and 5-aminoindazole (3) displayed efficient target protein interactions with a dose-dependent antinociceptive effect in the mice formalin test, which in the second phase was dependent on FAAH and TRPV1. No hepatotoxicity of the FAAH substrates transformed into TRPV1 activators was observed. While paracetamol attenuates pyrexia via inhibition of brain cyclooxygenase, its antinociceptive FAAH substrate 4-AP was not antipyretic, suggesting separate mechanisms for the antipyretic and antinociceptive effect of paracetamol. Furthermore, compound 3 reduced fever without a brain cyclooxygenase inhibitory action. The data support our view that analgesics and antipyretics without liver toxicity can be derived from paracetamol. Thus, research into the molecular actions of paracetamol could pave the way for the discovery of analgesics and antipyretics with a better benefit-to-risk ratio.

OBJECTIVES:Infections, cancer, and systemic inflammation elicit anorexia. Despite the medical significance of this phenomenon, the question of how peripheral inflammatory mediators affect the central regulation of food intake is incompletely understood. Therefore, we have investigated the sickness behavior induced by the prototypical inflammatory mediator IL-1β. METHODS:IL-1β was injected intravenously. To interfere with IL-1β signaling, we deleted the essential modulator of NF-κB signaling (Nemo) in astrocytes and tanycytes. RESULTS:Systemic IL-1β increased the activity of the transcription factor NF-κB in tanycytes of the mediobasal hypothalamus (MBH). By activating NF-κB signaling, IL-1β induced the expression of cyclooxygenase-2 (Cox-2) and stimulated the release of the anorexigenic prostaglandin E2 (PGE2) from tanycytes. When we deleted Nemo in astrocytes and tanycytes, the IL-1β-induced anorexia was alleviated whereas the fever response and lethargy response were unchanged. Similar results were obtained after the selective deletion of Nemo exclusively in tanycytes. CONCLUSIONS:Tanycytes form the brain barrier that mediates the anorexic effect of systemic inflammation in the hypothalamus.

We examined the role of brown adipose tissue (BAT) for fever and emotional stress-induced hyperthermia. Wild-type and uncoupling protein-1 (UCP-1) knockout mice were injected with lipopolysaccharide intraperitoneally or intravenously, or subjected to cage exchange, and body temperature monitored by telemetry. Both genotypes showed similar febrile responses to immune challenge and both displayed hyperthermia to emotional stress. Neither procedure resulted in the activation of BAT, such as the induction of UCP-1 or peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) mRNA, or reduced BAT weight and triglyceride content. In contrast, in mice injected with a β3 agonist, UCP-1 and PGC-1α were strongly induced, and BAT weight and triglyceride content reduced. Both lipopolysaccharide and the β3 agonist, and emotional stress, induced UCP-3 mRNA in skeletal muscle. A β3 antagonist did not attenuate lipopolysaccharide-induced fever, but augmented body temperature decrease and inhibited BAT activation when mice were exposed to cold. An α1 /α2b antagonist or a 5HT1A agonist, which inhibit vasoconstriction, abolished lipopolysaccharide-induced fever, but had no effect on emotional stress-induced hyperthermia. These findings demonstrate that in mice, UCP-1-mediated BAT thermogenesis does not take part in inflammation-induced fever, which is dependent on peripheral vasoconstriction, nor in stress-induced hyperthermia. However, both phenomena may involve UCP-3-mediated muscle thermogenesis.