Identification of therapeutic targets in a murine model of severe exertional heat stroke.

Exertional heat stroke (EHS) is a potentially lethal condition resulting from high core body temperatures (TC) in combination with a systemic inflammatory response syndrome (SIRS) with varying degrees of severity across victims, and limited understanding of the underlying mechanism(s). We established a mouse model of severe EHS to identify mechanisms of hyperthermia/inflammation that may be responsible for organ damage. Mice were forced to run on a motorized wheel in a 37.5°C chamber until loss of consciousness and were either removed immediately (exertional heat injury or EHI; TCMax = 42.4 ± 0.2°C) or remained in the chamber an additional 20 min (EHS; TCMax = 42.5 ± 0.4°C). Exercise control mice (ExC) experienced identical procedures to EHS at 25°C. At 3 h post-EHS, there was evidence for an immune/inflammatory response as elevated blood chemokine [interferon γ-induced protein 10 (IP-10), keratinocytes-derived chemokine (KC), macrophage inflammatory proteins (MIP-1α), MIP-1ß, MIP-2] and cytokine [granulocyte colony-stimulating factor (G-CSF), interleukins (IL-10), IL-6] levels peaked and were highest in EHS mice compared with EHI and ExC mice. Immunoblotting of organs susceptible to EHS damage indicated that several kinases were sensitive to stress associated with heat/inflammation and exercise; specifically, phosphorylation of liver c-Jun NH2-terminal kinase (JNK) at threonine 183/tyrosine 185 immediately (0 h) postheating related to heat illness severity. We have established a mouse EHS model, and JNK [or its downstream target(s)] could underlie EHS symptomatology, allowing the identification of molecular pathways or countermeasure targets to mitigate heat illness severity, enable complete recovery, and decrease overall EHS-related fatalities.

Prior viral illness increases heat stroke severity in mice.

NEW FINDINGS: What is the central question of this study? We hypothesized that prior illness would increase the susceptibility to and severity of heat stroke (HS). What is the main finding and its importance? We provide the first experimental evidence, using a mouse model of HS, that recent viral illness increases the severity of HS. Our data indicate that this effect is not attributable to the exacerbation of hyperthermia but is a consequence of ongoing coagulation and systemic inflammatory reactions. Our data suggest that measurement of platelets, cytokines and chemokines before heat exposure might be indicative of susceptibility to HS, with coagulation and inflammation being potential targets for intervention that could improve recovery. ABSTRACT: It is hypothesized that prior illness exacerbates heat stroke (HS) in otherwise healthy organisms by augmenting hyperthermia during heat exposure or deactivating cellular pathways that protect against organ injury. To test these hypotheses, we injected telemetered male C57BL/6J mice with lipopolysaccharide (LPS; 50 µg kg-1 i.p.) or polyinosinic:polycytidylic acid (PIC; 100 µg i.p.) as a bacterial or a viral mimic, respectively, with saline (SAL; equivalent volume) as a control. Mice recovered for 48 or 72 h before HS (maximal core temperature = 42.4°C). Platelet counts, cytokines, chemokines and organ injury were determined 48 or 72 h after injection (without heating) or at maximal core temperature and at 1 day of recovery from HS. In the absence of heat, PIC induced more robust signs of sickness and increased cytokines and chemokines (TNF-α, RANTES, IP-10 and MIP-1ß) at 48 h, which was not observed with LPS (P < 0.05). Responses of both groups recovered by 72 h, although low platelet counts persisted after PIC (P < 0.05). Heat-induced hyperthermia was similar among mice injected with SAL, LPS and PIC; however, PIC-injected mice displayed more severe responses during recovery from HS, with reduced survival (48 h, 70 versus 100%; P < 0.05), deeper and longer post-HS hypothermia, greater reductions in platelets, elevated RANTES, IP-10, IL-6 and TNF-α and greater duodenal injury (P < 0.05). By 72 h, survival from HS was no longer reduced in PIC-injected mice, although hypothermia, the reduction in platelets and elevated cytokines persisted. Our data indicate that prior illness exacerbates the severity of HS in the absence of signs of illness at the time of heat exposure and suggest that this is attributable to persistent coagulation and inflammatory reactions that might be targets for intervention to improve recovery.

The Role of Skeletal Muscles in Exertional Heat Stroke Pathophysiology.

The active participation of skeletal muscles is a unique characteristic of exertional heat stroke. Nevertheless, the only well-documented link between skeletal muscle activities and exertional heat stroke pathophysiology is the extensive muscle damage (e. g., rhabdomyolysis) and subsequent leakage of intramuscular content into the circulation of exertional heat stroke victims. Here, we will present and discuss rarely explored roles of skeletal muscles in the context of exertional heat stroke pathophysiology and recovery. This includes an overview of heat production that contributes to severe hyperthermia and the synthesis and secretion of bioactive molecules, such as cytokines, chemokines and acute phase proteins. These molecules can alter the overall inflammatory status from pro- to anti-inflammatory, affecting other organ systems and influencing recovery. The activation of innate immunity can determine whether a victim is ready to return to physical activity or experiences a prolonged convalescence. We also provide a brief discussion on whether heat acclimation can shift skeletal muscle secretory phenotype to prevent or aid recovery from exertional heat stroke. We conclude that skeletal muscles should be considered as a key organ system in exertional heat stroke pathophysiology.

Delayed metabolic dysfunction in myocardium following exertional heat stroke in mice.

KEY POINTS: Exposure to exertional heat stroke (EHS) is associated with increased risk of long-term cardiovascular disorders in humans. We demonstrate that in female mice, severe EHS results in metabolic changes in the myocardium, emerging only after 9-14 days. This was not observed in males that were symptom-limited at much lower exercise levels and heat loads compared to females. At 14 days of recovery in females, there were marked elevations in myocardial free fatty acids, ceramides and diacylglycerols, consistent with development of underlying cardiac abnormalities. Glycolysis shifted towards the pentose phosphate and glycerol-3-phosphate dehydrogenase pathways. There was evidence for oxidative stress, tissue injury and microscopic interstitial inflammation. The tricarboxylic acid cycle and nucleic acid metabolism pathways were also negatively affected. We conclude that exposure to EHS in female mice has the capacity to cause delayed metabolic disorders in the heart that could influence long-term health. ABSTRACT: Exposure to exertional heat stroke (EHS) is associated with a higher risk of long-term cardiovascular disease in humans. Whether this is a cause-and-effect relationship remains unknown. We studied the potential of EHS to contribute to the development of a 'silent' form of cardiovascular disease using a preclinical mouse model of EHS. Plasma and ventricular myocardial samples were collected over 14 days of recovery. Male and female C57bl/6J mice underwent forced wheel running for 1.5-3 h in a 37.5°C/40% relative humidity until symptom limitation, characterized by CNS dysfunction. They reached peak core temperatures of 42.2 ± 0.3°C. Females ran ∼40% longer, reaching ∼51% greater heat load. Myocardial and plasma samples (n = 8 per group) were obtained between 30 min and 14 days of recovery, analysed using metabolomics/lipidomics platforms and compared to exercise controls. The immediate recovery period revealed an acute energy substrate crisis from which both sexes recovered within 24 h. However, at 9-14 days, the myocardium of female mice developed marked elevations in free fatty acids, ceramides and diacylglycerols. Glycolytic and tricarboxylic acid cycle metabolites revealed bottlenecks in substrate flow, with build-up of intermediate metabolites consistent with oxidative stress and damage. Males exhibited only late stage reductions in acylcarnitines and elevations in acetylcarnitine. Histopathology at 14 days showed interstitial inflammation in the female hearts only. The results demonstrate that the myocardium of female mice is vulnerable to a slowly emerging metabolic disorder following EHS that may harbinger long-term cardiovascular complications. Lack of similar findings in males may reflect their lower heat exposure.

Cardiovascular and thermoregulatory dysregulation over 24 h following acute heat stress in rats.

The influences of severe heat stroke (HS) on cardiovascular function during recovery are incompletely understood. We hypothesized that HS would elicit a heart rate (HR) increase persisting through 24 h of recovery due to hemodynamic, thermoregulatory, and inflammatory events, necessitating tachycardia to support mean arterial pressure (MAP). Core temperature (Tc), HR, and MAP were measured via radiotelemetry in conscious male Fischer 344 rats (n = 22; 282.4 ± 3.5 g) during exposure to 37°C ambient temperature until a maximum Tc of 42.0°C, and during recovery at 20°C ambient temperature through 24 h. Rats were divided into Mild, Moderate, and Severe groups based on pathophysiology. HS rats exhibited hysteresis relative to Tc with HR higher for a given Tc during recovery compared with heating (P < 0.0001). "Reverse" hysteresis occurred in MAP with pressure during cooling lower than heating per degree Tc (P < 0.0001). Mild HS rats showed tachycardia [P < 0.01 vs. control (Con)] through 8 h of recovery, elevated MAP (P < 0.05 vs. Con) for the initial 5 h of recovery, with sustained hyperthermia (P < 0.05 vs. Con) through 24 h. Moderate HS rats showed significant tachycardia (P < 0.01 vs. Con), normal MAP (P > 0.05 vs. Con), and rebound hyperthermia from 4 to 24 h post-HS (P < 0.05 vs. Con). Severe HS rats showed tachycardia (P < 0.05 vs. Con), hypotension (P < 0.01 vs. Con), and hypothermia for 24 h (P < 0.05 vs. Con). Severe HS rats showed 14- and 12-fold increase in heart and liver inducible nitric oxide synthase expression, respectively. Hypotension and hypothermia in Severe HS rats was consistent with inducible nitric oxide synthase-mediated systemic vasodilation. These findings provide mechanistic insight into hemodynamic and thermoregulatory impairments during 24 h of HS recovery.

Point-of-care cardiac troponin test accurately predicts heat stroke severity in rats.

Heat stroke (HS) remains a significant public health concern. Despite the substantial threat posed by HS, there is still no field or clinical test of HS severity. We suggested previously that circulating cardiac troponin (cTnI) could serve as a robust biomarker of HS severity after heating. In the present study, we hypothesized that (cTnI) point-of-care test (ctPOC) could be used to predict severity and organ damage at the onset of HS. Conscious male Fischer 344 rats (n = 16) continuously monitored for heart rate (HR), blood pressure (BP), and core temperature (Tc) (radiotelemetry) were heated to maximum Tc (Tc,Max) of 41.9 ± 0.1°C and recovered undisturbed for 24 h at an ambient temperature of 20°C. Blood samples were taken at Tc,Max and 24 h after heat via submandibular bleed and analyzed on ctPOC test. POC cTnI band intensity was ranked using a simple four-point scale via two blinded observers and compared with cTnI levels measured by a clinical blood analyzer. Blood was also analyzed for biomarkers of systemic organ damage. HS severity, as previously defined using HR, BP, and recovery Tc profile during heat exposure, correlated strongly with cTnI (R(2) = 0.69) at Tc,Max. POC cTnI band intensity ranking accurately predicted cTnI levels (R(2) = 0.64) and HS severity (R(2) = 0.83). Five markers of systemic organ damage also correlated with ctPOC score (albumin, alanine aminotransferase, blood urea nitrogen, cholesterol, and total bilirubin; R(2) > 0.4). This suggests that cTnI POC tests can accurately determine HS severity and could serve as simple, portable, cost-effective HS field tests.

Patterns of gene expression associated with recovery and injury in heat-stressed rats.

BACKGROUND: The in vivo gene response associated with hyperthermia is poorly understood. Here, we perform a global, multiorgan characterization of the gene response to heat stress using an in vivo conscious rat model. RESULTS: We heated rats until implanted thermal probes indicated a maximal core temperature of 41.8°C (Tc,Max). We then compared transcriptomic profiles of liver, lung, kidney, and heart tissues harvested from groups of experimental animals at Tc,Max, 24 hours, and 48 hours after heat stress to time-matched controls kept at an ambient temperature. Cardiac histopathology at 48 hours supported persistent cardiac injury in three out of six animals. Microarray analysis identified 78 differentially expressed genes common to all four organs at Tc,Max. Self-organizing maps identified gene-specific signatures corresponding to protein-folding disorders in heat-stressed rats with histopathological evidence of cardiac injury at 48 hours. Quantitative proteomics analysis by iTRAQ (isobaric tag for relative and absolute quantitation) demonstrated that differential protein expression most closely matched the transcriptomic profile in heat-injured animals at 48 hours. Calculation of protein supersaturation scores supported an increased propensity of proteins to aggregate for proteins that were found to be changing in abundance at 24 hours and in animals with cardiac injury at 48 hours, suggesting a mechanistic association between protein misfolding and the heat-stress response. CONCLUSIONS: Pathway analyses at both the transcript and protein levels supported catastrophic deficits in energetics and cellular metabolism and activation of the unfolded protein response in heat-stressed rats with histopathological evidence of persistent heat injury, providing the basis for a systems-level physiological model of heat illness and recovery.

Alteration in circulating metabolites during and after heat stress in the conscious rat: potential biomarkers of exposure and organ-specific injury.

BACKGROUND: Heat illness is a debilitating and potentially life-threatening condition. Limited data are available to identify individuals with heat illness at greatest risk for organ damage. We recently described the transcriptomic and proteomic responses to heat injury and recovery in multiple organs in an in vivo model of conscious rats heated to a maximum core temperature of 41.8°C (Tc,Max). In this study, we examined changes in plasma metabolic networks at Tc,Max, 24, or 48 hours after the heat stress stimulus. RESULTS: Circulating metabolites were identified by gas chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. Bioinformatics analysis of the metabolomic data corroborated proteomics and transcriptomics data in the tissue at the pathway level, supporting modulations in metabolic networks including cell death or catabolism (pyrimidine and purine degradation, acetylation, sulfation, redox alterations and glutathione metabolism, and the urea cycle/creatinine metabolism), energetics (stasis in glycolysis and tricarboxylic acid cycle, ß-oxidation), cholesterol and nitric oxide metabolism, and bile acids. Hierarchical clustering identified 15 biochemicals that differentiated animals with histopathological evidence of cardiac injury at 48 hours from uninjured animals. The metabolic networks perturbed in the plasma corroborated the tissue proteomics and transcriptomics pathway data, supporting a model of irreversible cell death and decrements in energetics as key indicators of cardiac damage in response to heat stress. CONCLUSIONS: Integrating plasma metabolomics with tissue proteomics and transcriptomics supports a diagnostic approach to assessing individual susceptibility to organ injury and predicting recovery after heat stress.

Increased cytokine and chemokine gene expression in the CNS of mice during heat stroke recovery.

Heat stroke (HS) is characterized by a systemic inflammatory response syndrome (SIRS) consisting of profound core temperature (Tc) changes in mice. Encephalopathy is common at HS collapse, but inflammatory changes occurring in the brain during the SIRS remain unidentified. We determined the association between inflammatory gene expression changes in the brain with Tc disturbances during HS recovery in mice. Gene expression changes of heat shock protein (HSP)72, heme oxygenase (hmox1), cytokines (IL-1ß, IL-6, TNF-α), cyclooxygenase enzymes (COX-1, COX-2), chemokines (MCP-1, MIP-1α, MIP-1ß, CX3CR1), and glia activation markers (CD14, aif1, vimentin) were examined in the hypothalamus (HY) and hippocampus (HC) of control (Tc ∼ 36.0°C) and HS mice at Tc,Max (42.7°C), hypothermia depth (HD; 29.3 ± 0.4°C), and fever (37.8 ± 0.3°C). HSP72 (HY

Attenuated thermoregulatory, metabolic, and liver acute phase protein response to heat stroke in TNF receptor knockout mice.

Tumor necrosis factor (TNF) is considered an adverse mediator of heat stroke (HS) based on clinical studies showing high serum levels. However, soluble TNF receptors (sTNFR; TNF antagonists) were higher in survivors than nonsurvivors, and TNFR knockout (KO) mice showed a trend toward increased mortality, suggesting TNF has protective actions for recovery. We delineated TNF actions in HS by comparing thermoregulatory, metabolic, and inflammatory responses between B6129F2 (wild type, WT) and TNFR KO mice. Before heat exposure, TNFR KO mice showed ~0.4°C lower core temperature (T(c); radiotelemetry), ~10% lower metabolic rate (M(r); indirect calorimetry), and reduced plasma interleukin (IL)-1α and sIL-1RI than WT mice. KO mice selected warmer temperatures than WT mice in a gradient but remained hypothermic. In the calorimeter, both genotypes showed a similar heating rate, but TNFR KO maintained lower T(c) and M(r) than WT mice for a given heat exposure duration and required ~30 min longer to reach maximum T(c) (42.4°C). Plasma IL-6 increased at ~3 h of recovery in both genotypes, but KO mice showed a more robust sIL-6R response. Higher sIL-6R in the KO mice was associated with delayed liver p-STAT3 protein expression and attenuated serum amyloid A3 (SAA3) gene expression, suggesting the acute phase response (APR) was attenuated in these mice. Our data suggest that the absence of TNF signaling induced a regulated hypothermic state in the KO mice, TNF-IL-1 interactions may modulate T(c) and M(r) during homeostatic conditions, and TNF modulates the APR during HS recovery through interactions with the liver IL-6-STAT3 pathway of SAA3 regulation.

Ovariectomy aggravates the pathophysiological response to exertional heat stroke in mice.

Female mice have a greater capacity for exercising in the heat than male mice, reaching greater power output and longer times of heat exposure before succumbing to exertional heat stroke (EHS). Differences in body mass, size, or testosterone do not explain these distinct sex responses. Whether the ovaries could account for the superior exercise capacity in the heat in females remains unknown. Here, we determined the influence of ovariectomy (OVX) on exercise capacity in the heat, thermoregulation, intestinal damage, and heat shock response in a mouse EHS model. We performed bilateral OVX (n = 10) or sham (n = 8) surgeries in young adult (4 mo) female C57/BL6J mice. Upon recovery from surgeries, mice exercised on a forced wheel placed inside an environmental chamber set at 37.5 °C and 40% relative humidity until experiencing loss of consciousness (LOC). Terminal experiments were performed 3 h after LOC. OVX increased body mass by the time of EHS (sham = 3.8 ± 1.1, OVX = 8.3 ± 3.2 g, P < 0.05), resulted in shorter running distance (sham = 753 ± 189, OVX = 490 ± 87 m, P < 0.05), and shorter time to LOC (sham = 126.3 ± 21, OVX = 99.1 ± 19.8 min, P < 0.05). Histopathological assessment of the intestines revealed damage in the jejunum (sham = 0.2 ± 0.7, OVX = 2.1 ± 1.7 AU, P < 0.05) and ileum (sham = 0.3 ± 0.5, OVX = 1.8 ± 1.4 AU, P < 0.05). OVX increased mesenteric microvascular density (sham = 101 ± 25, OVX = 156 ± 66 10-2 mm/mm2, P < 0.05) and decreased concentration of circulatory heat shock protein 72 (HSP72) (sham = 26.7 ± 15.8, OVX = 10.3 ± 4.6 ng/mL, P < 0.05). No differences were observed in cytokines or chemokines between groups. Our findings indicate that OVX aggravates the pathophysiological response to EHS in mice.NEW & NOTEWORTHY Females outperform males in a mouse model of exertional heat stroke (EHS). Here, we show for the first time the impact of ovariectomy (OVX) on EHS pathophysiology. OVX resulted in a shorter exercise capacity in the heat, greater intestinal damage, and lower heat shock response following EHS.

Vancomycin modestly attenuates symptom severity during onset of and recovery from exertional heat stroke in mice.

Increased intestinal permeability during exertion and subsequent leakage of bacteria into circulation is hypothesized to accelerate exertional heat stroke (EHS) onset and/or exacerbate EHS severity. To provide proof of concept for this theory, we targeted intestinal microbiota via antibiotic prophylaxis and determined whether vancomycin would delay EHS onset and/or mitigate EHS severity and mortality rates using a mouse model of EHS. Mice were 1) designated as EHS or Exercise Control (ExC) and 2) given 7 days of vancomycin (VEHS, VExC) or untreated water (EHS, ExC) before EHS/Exercise. Following EHS/ExC, mice were euthanized immediately (0 h) or returned to their home cage (25°C) and euthanized after 3 h or 24 h. VEHS mice exhibited reduced abundance and altered composition of fecal bacteria (with notable decreases in genera within orders Clostridiales and Bacteroidales); increased water consumption, lower core temperature (TC) before and during heating (TCMax), lower circulating markers of organ damage and inflammation at 24 h; and reduced hepatic activation of stress pathways at 0 and 3 h compared with EHS mice. Vancomycin-induced alterations to the intestinal microbiota likely influenced EHS outcomes, but it is unconfirmed whether this is due to attenuated bacterial leakage into circulation or other (in)direct effects on physiology and behavior (e.g., decreased TC, increased water consumption). To our knowledge, this is the first study quantitating antibiotic effects in conscious/unanesthetized, exertional HS animals.NEW & NOTEWORTHY Vancomycin prophylaxis lowered core temperature before and during EHS, mitigated EHS-associated rise of hepatic biomarkers and cytokines/chemokines in circulation (particularly at 24 h), and corresponded to inhibited phosphorylation of hepatic c-Jun NH2-terminal kinase on Threonine 183/Tyrosine 185 at 0 and 3 h in conscious, unanesthetized mice. However, vancomycin also induced cecal enlargement suggesting its off-target effects could limit its utility against EHS.

Exertional heat stroke: pathophysiology and risk factors.

Exertional heat stroke, the third leading cause of mortality in athletes during physical activity, is the most severe manifestation of exertional heat illnesses. Exertional heat stroke is characterised by central nervous system dysfunction in people with hyperthermia during physical activity and can be influenced by environmental factors such as heatwaves, which extend the incidence of exertional heat stroke beyond athletics only. Epidemiological data indicate mortality rates of about 27%, and survivors display long term negative health consequences ranging from neurological to cardiovascular dysfunction. The pathophysiology of exertional heat stroke involves thermoregulatory and cardiovascular overload, resulting in severe hyperthermia and subsequent multiorgan injury due to a systemic inflammatory response syndrome and coagulopathy. Research about risk factors for exertional heat stroke remains limited, but dehydration, sex differences, ageing, body composition, and previous illness are thought to increase risk. Immediate cooling remains the most effective treatment strategy. In this review, we provide an overview of the current literature emphasising the pathophysiology and risk factors of exertional heat stroke, highlighting gaps in knowledge with the objective to stimulate future research.

Classic and exertional heatstroke.

In the past two decades, record-breaking heatwaves have caused an increasing number of heat-related deaths, including heatstroke, globally. Heatstroke is a heat illness characterized by the rapid rise of core body temperature above 40 °C and central nervous system dysfunction. It is categorized as classic when it results from passive exposure to extreme environmental heat and as exertional when it develops during strenuous exercise. Classic heatstroke occurs in epidemic form and contributes to 9-37% of heat-related fatalities during heatwaves. Exertional heatstroke sporadically affects predominantly young and healthy individuals. Under intensive care, mortality reaches 26.5% and 63.2% in exertional and classic heatstroke, respectively. Pathological studies disclose endothelial cell injury, inflammation, widespread thrombosis and bleeding in most organs. Survivors of heatstroke may experience long-term neurological and cardiovascular complications with a persistent risk of death. No specific therapy other than rapid cooling is available. Physiological and morphological factors contribute to the susceptibility to heatstroke. Future research should identify genetic factors that further describe individual heat illness risk and form the basis of precision-based public health response. Prioritizing research towards fundamental mechanism and diagnostic biomarker discovery is crucial for the design of specific management approaches.

Drug-Induced Hyperthermia Review.

Humans maintain core body temperature via a complicated system of physiologic mechanisms that counteract heat/cold fluctuations from metabolism, exertion, and the environment. Overextension of these mechanisms or disruption of body temperature homeostasis leads to bodily dysfunction, culminating in a syndrome analogous to exertional heat stroke (EHS). The inability of this thermoregulatory process to maintain the body temperature is caused by either thermal stress or certain drugs. EHS is a syndrome characterized by hyperthermia and the activation of systemic inflammation. Several drug-induced hyperthermic syndromes may resemble EHS and share common mechanisms. The purpose of this article is to review the current literature and compare exertional heat stroke (EHS) to three of the most widely studied drug-induced hyperthermic syndromes: malignant hyperthermia (MH), neuroleptic malignant syndrome (NMS), and serotonin syndrome (SS). Drugs and drug classes that have been implicated in these conditions include amphetamines, diuretics, cocaine, antipsychotics, metoclopramide, selective serotonin reuptake inhibitors (SSRIs), tricyclic antidepressants (TCAs), and many more. Observations suggest that severe or fulminant cases of drug-induced hyperthermia may evolve into an inflammatory syndrome best described as heat stroke. Their underlying mechanisms, symptoms, and treatment approaches will be reviewed to assist in accurate diagnosis, which will impact the management of potentially life-threatening complications.

Tissue and circulating expression of IL-1 family members following heat stroke.

Interleukin-1 (IL-1) is thought to have a significant role in the pathophysiology of heat stroke (HS), although little is known regarding the actions or expression patterns of the IL-1 family. This study tested the hypotheses that following HS IL-1 family gene expression is dynamic, while loss of IL-1 signaling enhances recovery. IL-1 family expression was determined in plasma, spleen, and liver from C57BL/6J mice (n=24 control, n=20 HS) at maximum core temperature (Tc,Max), hypothermia, and 24 h post-HS (24 h). Soluble IL-1 receptor subtype I (sIL-1RI) protein expression peaked at 24 h (14,659.01±2,016.28 pg/ml, P<0.05), while sIL-1RII peaked at hypothermia (19,099.30±1,177.07 pg/ml). IL-1α gene expression in the spleen (ninefold) and liver (fourfold) along with IL-1RI (threefold spleen and fivefold liver) were maximal at hypothermia. Spleen IL-1ß gene expression peaked at Tc,Max (fourfold) but at hypothermia (fourfold) in liver. Gene expression of the IL-1 family member IL-18 peaked (2.5-fold) at Tc,Max but was similar at all other time points. Subsequent studies revealed that despite accruing a greater heating area (298±16 vs. 247±13°C·min, P<0.05), IL-1RI knockout (KO) mice (n=14) showed an attenuated hypothermia depth (28.5±0.2 vs. 27.3±0.5°C, P<0.05) and duration (675±82 vs. 1,283±390 min, P<0.05) with a higher 24 h Tc (36.9 vs. 34.1°C, P<0.05) compared with C57BL/6J mice (n=8). The current results demonstrate that following HS IL-1 family gene expression is altered and IL-1RI KO mice display Tc responses consistent with a more rapid recovery.

Impact of successive exertional heat injuries on thermoregulatory and systemic inflammatory responses in mice.

The purpose of the study was to determine if repeated exertional heat injuries (EHIs) worsen the inflammatory response. We assessed the impact of a single EHI bout (EHI0) or two separate EHI episodes separated by 1 (EHI1), 3 (EHI3), and 7 (EHI7) days in male C57BL/6J mice (n = 236). To induce EHI, mice underwent a forced running protocol until loss of consciousness or core temperature reached ≥ 42.7°C. Blood and tissue samples were obtained 30 min, 3 h, 1 day, or 7 days after the EHI. We observed that mice undergoing repeated EHI (EHI1, EHI3, and EHI7) had longer running distances before collapse (∼528 m), tolerated higher core temperatures (∼0.18°C higher) before collapse, and had higher minimum core temperature (indicative of injury severity) during recovery relative to EHI0 group (∼2.18°C higher; all P < 0.05). Heat resilience was most pronounced when latency was shortest between EHI episodes (i.e., thermal load and running duration highest in EHI1), suggesting the response diminishes with longer recoveries between EHI events. Furthermore, mice experiencing a second EHI exhibited increased serum and liver HSP70, and lower corticosterone, FABP2, MIP-1ß, MIP-2, and IP-10 relative to mice experiencing a single EHI typically at 30 min to 3 h after EHI. Our findings indicate that an EHI event may initiate some adaptive processes that provide acute heat resilience to subsequent EHI conditions. NEW & NOTEWORTHY Mice undergoing repeated exertional heat injuries, within 1 wk of an initial heat injury, appear to have some protective adaptations. During the second exertional heat injury, mice were able to run longer and sustain higher body temperatures before collapse. Despite this, the mice undergoing a second exertional heat injury were more resilient to the heat as evidenced by attenuated minimum body temperature, higher HPS70 (serum and liver), lower corticosterone, and lower FABP2.

Thermoregulatory, behavioral, and metabolic responses to heatstroke in a conscious mouse model.

The typical core temperature (T(c)) profile displayed during heatstroke (HS) recovery consists of initial hypothermia followed by delayed hyperthermia. Anecdotal observations led to the conclusion that these T(c) responses represent thermoregulatory dysfunction as a result of brain damage. We hypothesized that these T(c) responses are mediated by a change in the temperature setpoint. T(c) (+/- 0.1 degrees C; radiotelemetry) of male C57BL/6J mice was monitored while they were housed in a temperature gradient with ambient temperature (T(a)) range of 20-39 degrees C to monitor behaviorally selected T(a) (T(s)) or an indirect calorimeter (T(a) = 25 degrees C) to monitor metabolism (V(O(2))) and calculate respiratory exchange ratio (RER). Responses to mild and severe HS (thermal area 249.6 +/- 18.9 vs. 299.4 +/- 19.3 degrees C.min, respectively) were examined through 48 h of recovery. An initial hypothermia following mild HS was associated with warm T(s) (approximately 32 degrees C), approximately 35% V(O(2)) decrease, and RER approximately 0.71 that indicated reliance on fatty acid oxidation. After 24 h, mild HS mice developed hyperthermia associated with warm T(s) (approximately 32 degrees C), approximately 20% V(O(2)) increase, and RER approximately 0.85. Severe HS mice appeared poikilothermic-like in the temperature gradient with T(c) similar to T(s) (approximately 20 degrees C), and these mice failed to recover from hypothermia and develop delayed hyperthermia. Cellular damage (hematoxylin and eosin staining) was undetectable in the hypothalamus or other brain regions in severe HS mice. Overall, decreases and increases in T(c) were associated with behavioral and autonomic thermoeffectors that suggest HS elicits anapyrexia and fever, respectively. Taken together, T(c) responses of mild and severe HS mice suggest a need for reinterpretation of the mechanisms of thermoregulatory control during recovery.

Overlapping Mechanisms of Exertional Heat Stroke and Malignant Hyperthermia: Evidence vs. Conjecture.

Exertional heat stroke (EHS) and malignant hyperthermia (MH) are life-threatening conditions, triggered by different environmental stimuli that share several clinical symptoms and pathophysiological features. EHS manifests during physical activity normally, but not always, in hot and humid environments. MH manifests during exposure to haloalkane anesthetics or succinylcholine, which leads to a rapid, unregulated release of calcium (Ca2+) within the skeletal muscles inducing a positive-feedback loop within the excitation-contraction coupling mechanism that culminates in heat stroke-like symptoms, if not rapidly recognized and treated. Rare cases of awake MH, independent of anesthesia exposure, occur during exercise and heat stress. It has been suggested that EHS and MH are mediated by similar mechanisms, including mutations in Ca2+ regulatory channels within the skeletal muscle. Rapid cooling, which is the most effective treatment for EHS, is ineffective as an MH treatment; rather, a ryanodine receptor antagonist drug, dantrolene sodium (DS), is administered to the victim to prevent further muscle contractions and hyperthermia. Whether DS can be an effective treatment for EHS victims remains uncertain. In the last decade, multiple reports have suggested a number of mechanistic links between EHS and MH. Here, we discuss aspects related to the pathophysiology, incidence, diagnosis and treatment. Furthermore, we present evidence regarding potential overlapping mechanisms between EHS and MH and explore current knowledge to establish what is supported by evidence or a lack thereof (i.e. conjecture).