Heat stress-induced neuroinflammation and aberration in monoamine levels in hypothalamus are associated with temperature dysregulation

Highlights

• Baroreflex responsiveness is intact under Moderate HS and early stages of Severe HS.

• HTH elicits normal neurotransmitter release in response to Moderate HS.

• Severe HS induces neuroinflammation in HTH causing Monoamines & Glu imbalance culminating in thermoregulatory dysfunction.

Abstract

Heat Stress (HS) induces diverse pathophysiological changes, which include brain ischemia, oxidative stress and neuronal damage. The present study was undertaken with the objective to ascertain whether neuroinflammation in Hypothalamus (HTH) caused under HS affects monoamine levels and hence, its physiological role in thermoregulation. Rats were exposed to HS in a heat simulation environmental chamber (Ambient temperature, Ta = 45 ± 0.5 °C and Relative Humidity, RH = 30 ± 10%) with real-time measurement of core temperature (Tc) and skin temperature (Ts). Animals were divided into two subgroups: Moderate HS (MHS) (Tc = 40 °C) and Severe HS (SHS)/Heat stroke (Tc = 42 °C). Rats with MHS showed an increase in Mean Arterial Pressure (MAP) and Heart Rate (HR) while fall in MAP and rise in HR was observed in rats with SHS. In addition, oxidative stress and an increase in pyknotic neurons were observed in HTH. High levels of Adrenocorticotropic-hormone (ACTH), Epinephrine (EPI), Norepinephrine (NE) and Dopamine (DA) in the systemic circulation and progressive increase in EPI and DA levels in HTH were recorded after the thermal insult. Moreover, a substantial increase in Glutamate (Glu) level was observed in HTH as well as in systemic circulation of heat stroke rats. We found a rise in NE whereas a fall in Serotonin (5-HT) level in HTH at MHS, without perturbing inflammatory mediators. However, rats with SHS exhibited significant elevations in NF-kB, IL-1β, COX2, GFAP and Iba1 protein expression in HTH. In conclusion, the data suggest that SHS induces neuroinflammation in HTH, which is associated with monoamines and Glu imbalances, leading to thermoregulatory disruption.

Introduction

Exposure to high ambient temperature is a natural hazard with an increasing rate of mortality across the globe. The armed forces personnel deployed in desert areas as well as the farmers, laborers and fire-fighters are often exposed to extreme environmental HS which affects their physiology and performance. The problem continues to increase in the degree of severity as the global temperature is rising. Consequently, heat stroke-related deaths have increased worldwide (Petoukhov et al., 2013, Horton et al., 2016, Li et al., 2016). Recently, Mann et al. described the influence of anthropogenic climate change on extreme weather events (Mann et al., 2017). Heat stroke is a life-threatening pathological condition, which is characterized by hyperthermia associated with systemic inflammatory response that leads to multi organ dysfunction syndrome (MODS), in which central nervous system (CNS) maladies prevail (Bouchama and Knochel, 2002, Chen et al., 2013, Singh et al., 2013, Leon and Bouchama, 2015). Studies have also shown that 30% of heat stroke survivors experience disability and neurological dysfunction even after cooling of the whole body (Dematte et al., 1998, Argaud et al., 2007, Wallace et al., 2007). It is evident from the reports that brain ischemia, inflammation and neuronal damage are the main causes of heat stroke, rather than body hyperthermia (Shih et al., 1984, Chen et al., 2006, Shen et al., 2008, Yang et al., 2010). So, there is a pressing need for the design of diagnostic and therapeutic modalities for heat stroke.

Heat exposure stimulates a myriad of biological stress events (McEwen, 2005) and protein expression in different brain regions including HTH. The preoptic area (POA), in and near the rostral HTH, acts as a coordinating center and strongly influences each of the lower effector areas. It is composed of neurons that are sensitive to minute modulations in hypothalamic or core temperature. The POA elicits thermoregulatory responses to maintain homeostasis for both internal and environmental thermal conditions (Boulant, 2000). Hypothalamic paraventricular nucleus (PVN) has been described as the ‘autonomic master controller’ and coordinates critical physiological responses by regulating the hypothalamo-pituitary-adrenocortical (HPA) axis (Smith and Vale, 2006, Ferguson et al., 2008). Michel et al. have proposed that decreased heat tolerance is associated with HPA axis impairment (Michel et al., 2007). Furthermore, HPA axis mobilization is accompanied by stimulation of the sympathetic system, as suggested by an increase in plasma NE and EPI levels (Gisolfi et al., 1991, Kregel et al., 1991) and an increased release of hypothalamic DA and 5-HT (Lin, 1997) upon heat exposure. Reports also indicate that predisposition to certain chronic systemic inflammatory conditions like diabetes, Chronic Obstructive Pulmonary Disease (COPD) and drug abuse reduce heat tolerance (Schmidt et al., 1999, Sevenoaks and Stockley, 2006, Leon, 2007, Chen et al., 2013). Moreover, a single episode of hyperthermia may cause short-term neurological dysfunction, which may become prolonged or permanent (Walter and Carraretto, 2016).

Hence, the studies aimed at unravelling the effect of HS on monoamines release in HTH are only sparse and investigating the link between HS-induced neuroinflammation and monoamine imbalance might provide insights into developing strategies to combat heat stroke. Therefore, we hypothesized that SHS induces neuroinflammation in HTH which is followed by an aberration in the release of monoamines, leading to temperature dysregulation. To test the hypothesis, we have designed a multi-parametric study in which the heat-stressed rats have been divided into two subgroups: subgroup 1, where the rats attained Tc of 40 °C, MHS (Liu et al., 2011) and subgroup 2 with Tc of 42 °C, SHS (Kao et al., 1994, Yang and Lin, 2002, Lee et al., 2005, Yang et al., 2010, Stallings et al., 2014, Audet et al., 2016). The rationale behind incorporating the two subgroups can be explained by the fact that Tc around 40 °C is considered as moderate heat and non-lethal to the rats (Hubbard et al., 1976) and Tc >41 °C induces heat stroke (Leon, 2006, Yang et al., 2010, Audet et al., 2016, Bruchim et al., 2016). Although there was a difference of 2 °C in Tc between the experimental groups, it resulted in huge difference at the physiological, cellular and molecular level. Thus, the two subgroups provide a clear demarcation between the normal physiology and pathophysiology associated with the respective groups.

In the present study, we measured systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR) after heat exposure. EPI, NE and 5-HT levels along with the ischemic markers (Glu and DA) in HTH were estimated after the thermal insult. In addition, stress hormones like ACTH along with monoamines and Glu in systemic circulation were assayed to examine the effect of HS, as the circulatory dysfunction is a common feature of heat stroke (Leon and Bouchama, 2015). Oxidative stress markers such as reactive oxygen species (ROS), Malondialdehyde (MDA), Protein oxidation, oxidized glutathione/reduced glutathione (GSSG/GSH) ratio, Neuronal damage (Cresyl violet staining) and the expression levels of inflammatory mediators like Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-kB p65), Interleukin-1 beta (IL-1β), Cyclooxygenase-2 (COX2), Glial Fibrillary Acidic Protein (GFAP), Ionized calcium-binding adapter molecule 1 (Iba 1) were estimated in HTH of rats after thermal insult.