Inflammation Emerges as a Sensitizer of a Heat-Responsive Cation Channel

Abstract

The Impact of Inflammation on Thermal Hyperpnea: Relevance for Heat Stress and Febrile Seizures.

Barrett KT, Roy A, Ebdalla A, Pittman QJ, Wilson RJA, Scantlebury MH. Am J Respir Cell Mol Biol. 2024 Aug;71(2):195–206. doi: 10.1165/rcmb.2023-0451OC. PMID: 38597725; PMCID: PMC11299082.

Extreme heat caused by climate change is increasing the transmission of infectious diseases, resulting in a sharp rise in heat-related illness and mortality. Understanding the mechanistic link between heat, inflammation, and disease is thus important for public health. Thermal hyperpnea, and consequent respiratory alkalosis, is crucial in febrile seizures and convulsions induced by heat stress in humans. Here, we address what causes thermal hyperpnea in neonates and how it is affected by inflammation. Transient receptor potential cation channel subfamily V member 1 (TRPV1), a heat-activated channel, is sensitized by inflammation and modulates breathing and thus may play a key role. To investigate whether inflammatory sensitization of TRPV1 modifies neonatal ventilatory responses to heat stress, leading to respiratory alkalosis and an increased susceptibility to hyperthermic seizures, we treated neonatal rats with bacterial LPS, and breathing, arterial pH, in vitro vagus nerve activity, and seizure susceptibility were assessed during heat stress in the presence or absence of a TRPV1 antagonist (AMG-9810) or shRNA-mediated TRPV1 suppression. LPS-induced inflammatory preconditioning lowered the threshold temperature and latency of hyperthermic seizures. This was accompanied by increased tidal volume, minute ventilation, expired CO₂, and arterial pH (alkalosis). LPS exposure also elevated vagal spiking and intracellular calcium concentrations in response to hyperthermia. TRPV1 inhibition with AMG-9810 or shRNA reduced the LPS-induced susceptibility to hyperthermic seizures and altered the breathing pattern too fast shallow breaths (tachypnea), making each breathless efficient and restoring arterial pH. These results indicate that inflammation exacerbates thermal hyperpnea-induced respiratory alkalosis associated with increased susceptibility to hyperthermic seizures, primarily mediated by TRPV1 localized to vagus neurons.

Commentary

Febrile seizure (FS) is a prevalent neurologic condition in children. It is characterized by seizures occurring with fever, typically in children between six months and five years of age. FS represents the most common type of seizure in patients without an underlying chronic neurologic disorder. ¹ The causes of FS are multifactorial and heterogeneous, but the underlying pathophysiology of how elevated body temperature induces seizures is unresolved.

In addition to convulsions, fever-associated hyperventilation can occur in children who experience FS. Hyperventilation and thermal hyperpnea can sometimes lead to respiratory alkalosis, ² but the underlying relationships between fever, hyperventilation, alkalosis, and the resultant seizures remain ill-defined. In experimental rodent models, FS can be induced in the absence of inflammation. ³ However, most clinical cases of FS occur in an elevated systemic inflammatory context brought on by a viral infection. Thus, inflammation is an invariant feature of the clinical FS presentation.

Transient receptor potential cation channel subfamily V member 1 (TRPV1) is a nonselective cation channel highly permeable to calcium cations. TRPV1 channels are activated by capsaicin or heat. They are expressed in immune cells and central and peripheral nervous tissues that regulate the respiratory response to elevated temperatures, including the carotid sinus and vagus nerves. ⁴ Previous work by Barrett et al ⁵ identified vagal TRPV1 engagement in increased FS susceptibility in postnatal day (P) 10 rats, providing an anatomical and pharmacological target for drug development. Studies performed in tissue cultures indicate the ambient temperature activation threshold for TRPV1 is ∼45°C,^4,6 which is higher than patient temperatures typical for FS. Thus, if TRPV1 activation plays a prominent role in FS, the channel would need to be sensitized to lower the activation threshold. One potent TRPV1 sensitizer is inflammation. These considerations led Barrett et al ⁷ to hypothesize that chronic inflammation, acting through TRPV1 activation, enhances hyperthermia-induced respiratory alkalosis and propensity to seize.

A chronic inflammatory condition was created in P10 rats using repeated low doses of lipopolysaccharide (LPS), a component of the cell wall of gram-negative bacteria. P10 rats subject to inflammatory preconditioning and FS showed altered inflammatory profiles in serum, reduced seizure threshold, and latency to first seizures, which agrees with previous reports. ⁸ The FS susceptibility was associated with thermal hyperpnea that resulted in respiratory alkalosis. After inflammation, the vagal response to heat was quicker, more enduring, and engaged at lower temperatures. Notably, intracellular calcium measurements supported the role of TRPV1 in breathing control in vagal nodes.

The critical experiments used a potent and selective antagonist for TRPV1 previously known to inhibit capsaicin-induced eye-wiping and hyperalgesia. ⁹ Administration of the TRPV1 antagonist, AMG-9810, rescued LPS-induced depression in FS threshold and seizure latency, thermal hypernea, and respiratory alkalosis. Thus, TRPV1 antagonism reversed the effect of inflammatory preconditioning on FS susceptibility. Complementary experiments then employed shRNA to knock down vagal TRPV1. Trpv1 mRNA levels were decreased by ∼15%, and this reduction was associated with suppressed effects of LPS-induced inflammation on neuronal activity in response to heat and FS latency. The overall conclusion from this study is that TRPV1 is a mediator of inflammation-induced respiratory alkalosis and associated susceptibility to FS. The current experimental design assessed TRPV1 sensitization within hours of the final LPS injection. It would be clinically relevant to determine if LPS-induced TRPV1 sensitization continues after the initial inflammatory event resolves.

Utilization of both male and female rats from multiple litters, preliminary and thorough characterization of the consequences of LPS administration in neonatal rats, and appropriate statistical analyses greatly enhanced the rigor and reproducibility of the study. The manuscript would be improved by reporting more drug dosing details, such as timing and vehicle, drug occupancy–efficacy relationships, and pharmacokinetic/pharmacodynamic data appropriate for the drug, vehicle, and route of administration.

To date, several TRPV1 antagonists have entered clinical trials to treat pain. One antagonist showed promise as an effective analgesic in preclinical animal studies, but phase 1 clinical trials were halted due to the emergence of hyperthermia as a side effect.^10,11 Another TRPV1 antagonist promoted skin tumors in mice. ¹² Advancement of TRPV1 as a therapeutic target in a clinical setting will likely require further preclinical development. Rodents pretreated with bacterial LPS have a greater susceptibility to chemoconvulsant-induced seizures and resultant inflammation and neuronal damage. However, viral, not bacterial, infections are the primary cause of fever in children with FS. The study by Barrett et al used LPS, which is one limitation of the current study that might impact the translatability of the treatment of FS in humans.

Another possible confound of this study is the use of P10 rats. Rats are altricial; thus nervous development for P10 rats roughly corresponds to the cerebral development stage of a newborn human. ¹³ To better understand the impact of TRPV1 activation in FS and enhance the translation impact, it would be informative to repeat the study but with P20 rats that correspond to 2–3-year-old humans.

In summary, this study builds momentum toward introducing TRPV1 receptor antagonists as adjunctive therapy for FS. The paper adds a pharmacologic and genetic case to this clinical use and thus provides further support from this group's work in similar preclinical models. The use of TRPV1 antagonists in the clinic still requires further investigation. Early-life inflammation may permanently sensitive TRPV1 channels, which could adversely impact pediatric populations experiencing recurrent infections. Cumulative TRPV1 activation could heighten seizure risk into later life. Trpv1 knockout exacerbates hyperthermic seizures in some mouse strains. ¹⁴ TRPV1 inhibition exhibits divergent and context-dependent outcomes. Limiting TRPV1 inhibition to periods of fever spikes could be a viable therapeutic strategy to mitigate the potential adverse effects of more prolonged TRPV1 inhibition. Fever-reducing and cooling strategies may be more effective and met with fewer challenges than TRPV1 inhibition as a therapy. Nevertheless, future well-designed studies employing viral-based inflammation, different ages of models, and additional species would provide important insights and rationale for the continued development of TRPV1 as a target.