Inhibition of microglial activation with minocycline at the intrathecal level attenuates sympathoexcitatory and proarrhythmogenic changes in rats with chronic temporal lobe epilepsy
Introduction
Epilepsy is a chronic brain disorder characterized by spontaneous recurrent seizures and carries a risk of sudden death that is 15–20 times higher than in normal population (Ficker et al., 1998, Nilsson et al., 1999, Eastaugh et al., 2015). Epilepsy affects about 50 million people worldwide (WHO, 2005); seizures can range from brief, barely noticeable loss of attention to major convulsions that affect the entire neuraxis. Epilepsy is associated with changes in autonomic functions, such as sympathovagal imbalance, sympathetic reflex dysfunction, tachycardia with concomitant arrhythmia or bradycardia with associated apnea (Dütsch et al., 2006, Bateman et al., 2008, Ponnusamy et al., 2012, Massey et al., 2014, Powell et al., 2014b, Bhandare et al., 2015, Bhandare et al., 2016a). Seizure-associated autonomic cardiorespiratory changes are well-documented and are thought to play an important role in a mechanism of sudden unexpected death in epilepsy (SUDEP) (Nei et al., 2004, Dlouhy et al., 2015). Interictal autonomic changes are also seen in patients with chronic epilepsy (Ansakorpi et al., 2000, Berilgen et al., 2004, Müngen et al., 2010, Lotufo et al., 2012). Nevertheless, the neuronal mechanisms causing autonomic cardiorespiratory dysfunction during chronic epilepsy are unknown.
Pituitary adenylate cyclase-activating polypeptide (PACAP), a 38 amino acid pleiotropic neuropeptide, produce neuroprotective effects (Shioda et al., 1998, Ohtaki et al., 2006, Bhandare et al., 2015) that are partly mediated through its action on microglia (Wada et al., 2013, Brifault et al., 2015). PACAP and microglia have a protective effect on sympathetic preganglionic neurons at the intermediolateral cell column (IML), where they ameliorate the sympathoexcitatory effect of acute seizures (Bhandare et al., 2015). During acute seizures, PACAP and microglia act on presympathetic rostral ventrolateral medulla (RVLM) neurons in the brainstem to promote proarrhythmogenic changes, but not sympathoexcitation (Bhandare et al., 2016a). In many cardiovascular autonomic nuclei PACAP is pressor and sympathoexcitatory (Farnham et al., 2008, Farnham et al., 2011, Inglott et al., 2011) and changes baroreflex response in trout (Lancien et al., 2011) but not in rats (Farnham et al., 2012). PACAP expression is increased in central autonomic nuclei (paraventricular nucleus) after kainic acid (KA)-induced seizures in rats (Nomura et al., 2000). Secondly, seizures produce microglial activation, and neuroinflammation in patients and animal models (Beach et al., 1995, Shapiro et al., 2008, Eyo et al., 2014), which persist for many years during chronic epilepsy (Beach et al., 1995, Papageorgiou et al., 2015). Microglia can be pro- or anti-inflammatory in animal models of temporal lobe epilepsy (TLE) (Shapiro et al., 2008, Mirrione et al., 2010, Vinet et al., 2012, Devinsky et al., 2013). Although the pro- or anti-inflammatory state of activated microglia is a topic of debate, there is strong support for their dual role (Hanisch and Kettenmann, 2007). Short-term microglial activation is considered beneficial (Mirrione et al., 2010, Vinet et al., 2012, Szalay et al., 2016), whereas chronic microglial activation is deleterious, and produces a damaging response to injury (Qin et al., 2007, Loane et al., 2014, Olmos-Alonso et al., 2016). During KA-induced acute seizures, spinal microglia have a protective effect on sympathetic preganglionic neurons (Bhandare et al., 2015), however, their role in chronic epilepsy is not known.
Thus, the aims of this study were to identify the role of PACAP and microglia in the spinal cord, during chronic epilepsy, in the regulation of central autonomic cardiovascular activity. To achieve these aims we used a model of acquired epilepsy in rats that manifest spontaneous seizures and many features of acquired epilepsy in humans- the KA-induced post-status epilepticus (post-SE) model (Morimoto et al., 2004, Powell et al., 2008, Jupp et al., 2012). The effect of intrathecal (IT) infusion of the PACAP antagonist, PACAP(6–38), and the microglial antagonist, minocycline, on sympathetic activity, cardiovascular reflex responses, and the electrocardiogram (ECG) were analyzed in chronically epileptic and control rats. Microglial morphology and their phenotype in the vicinity of RVLM neurons were analyzed with immunohistochemistry in epileptic and control rats.
Section snippets
Animals
The animal usage and protocols were in accordance with the Australian code of practice for the care and use of animals for scientific purposes. The protocols were approved by the Animal Care, and Ethics Committee of Macquarie University, The University of Melbourne, and the Sydney Local Health District. The epilepsy surgery and procedures were performed under isoflurane anesthesia on 17–19-week-old adult non-epileptic control (n = 9), and post-SE (n = 15) male Wistar rats, whereas electrophysiology
Development of spontaneous recurrent seizures in post-SE rats
A typical spontaneous recurrent seizure with transition into ictal period is shown in Fig. 2, which is characterized by high-amplitude and showed a clear new pattern of tracing. Video-EEG-ECG recordings confirmed that 9 weeks after induction of KA-induced SE, almost all rats developed spontaneous recurrent seizures with 0.44 ± 0.07 seizures per day (range 0–3) and with duration of 19.7 ± 3.3 s per day (Table 1). The seizure frequency and scores are highly variable across the post-SE rats as shown in
Discussion
The study provides direct evidence that microglia play a role in mediating increased SNA, and arrhythmogenic cardiac electrophysiological changes in chronic epileptic rats. First, spontaneous seizures cause severe tachycardia with prolongation of QT interval that persisted for more than 1 h after the onset of a seizure. Secondly, antagonism of microglial activation, but not PACAP, in the spinal cord significantly reduces the SNA and seizure-induced prolongation of QTc interval in epileptic rats.
Conflict of interest
Authors have no conflicts of interest to declare.
Acknowledgments
Work in the Authors’ laboratory is supported by grants from the Australian Research Council (Discovery Early Career Researcher Award; DE120100992), National Health and Medical Research Council of Australia (1024489, 1065485, 1082215 and 1082215). AMB and KK are supported by international Macquarie University Research Excellence Scholarships (2012219 and 2012112), The Heart Research Institute (HRI) and The University of Sydney.
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2021, Autonomic Neuroscience: Basic and ClinicalCitation Excerpt :The primed animals go on to develop spontaneous recurrent seizures, which persist for weeks and may be lethal. These post SE models develop cardiac changes that could pre-dispose to arrhythmias (Auzmendi et al., 2018; Bhandare et al., 2017; Brewster et al., 2016; Little and Bealer, 2012; Powell et al., 2014; Sharma et al., 2020). Interpretation of results may, however, be complicated by the initial period of SE as well as by direct actions on peripheral or central (e.g. brainstem) sites, which can induce cardiac and respiratory changes independently of seizures.
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2021, NeuropharmacologyCitation Excerpt :Cardiovascular and respiratory chemoreflex responses to insufficient O2 and excessive CO2 are likely to be a critical lifesaving response during the postictal period (Massey et al., 2014). Interestingly, rats with chronic epilepsy due to kainic acid induced status epilepticus display normal chemoreflex responses to hypoxic and hypercapnic stimuli (Bhandare et al., 2017). It is conceivable that recent seizure activity, but not chronic epilepsy itself, causes chemoreflex disruption.
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2020, Brain, Behavior, and ImmunityCitation Excerpt :Consequently, our results suggest that microglia are overall protective and play a beneficial role in controlling KA-induced seizures and epilepsy. It is increasingly suggested that microglia have opposing roles in seizures, being beneficial during the acute phase and deleterious during the chronic phase (Bhandare et al., 2017). However, contrary to some reports that showed preventing microglia activity during that chronic phase was beneficial (Bhandare et al., 2017; Wang et al., 2015), we found microglia depletion increased the frequency of SRS and seizure duration.