Research paper
Vulnerability of cholecystokinin-expressing GABAergic interneurons in the unilateral intrahippocampal kainate mouse model of temporal lobe epilepsy

https://doi.org/10.1016/j.expneurol.2021.113724Get rights and content

Highlights

  • Unilateral dorsal IHK mouse model of TLE reproduced key properties of human TLE.

  • The IHK model reduced CCK+ interneurons in the ventral hippocampus.

  • The IHK model reduced the number of CCKBC boutons in the ventral hippocampus.

  • The IHK model didn't alter intrinsic and synaptic properties of surviving CCKBCs.

  • The IHK model reduced overall CCK+ interneuron-mediated inhibition.

Abstract

Temporal lobe epilepsy (TLE) is characterized by recurrent spontaneous seizures and behavioral comorbidities. Reduced hippocampal theta oscillations and hyperexcitability that contribute to cognitive deficits and spontaneous seizures are present beyond the sclerotic hippocampus in TLE. However, the mechanisms underlying compromised network oscillations and hyperexcitability observed in circuits remote from the sclerotic hippocampus are largely unknown. Cholecystokinin (CCK)-expressing basket cells (CCKBCs) critically participate in hippocampal theta rhythmogenesis, and regulate neuronal excitability. Thus, we examined whether CCKBCs were vulnerable in nonsclerotic regions of the ventral hippocampus remote from dorsal sclerotic hippocampus using the intrahippocampal kainate (IHK) mouse model of TLE, targeting unilateral dorsal hippocampus. We found a decrease in the number of CCK+ interneurons in ipsilateral ventral CA1 regions from epileptic mice compared to those from sham controls. We also found that the number of boutons from CCK+ interneurons was reduced in the stratum pyramidale, but not in other CA1 layers, of ipsilateral hippocampus in epileptic mice, suggesting that CCKBCs are vulnerable. Electrical recordings showed that synaptic connectivity and strength from surviving CCKBCs to CA1 pyramidal cells (PCs) were similar between epileptic mice and sham controls. In agreement with reduced CCKBC number in TLE, electrical recordings revealed a significant reduction in amplitude and frequency of IPSCs in CA1 PCs evoked by carbachol (commonly used to excite CCK+ interneurons) in ventral CA1 regions from epileptic mice versus sham controls. These findings suggest that loss of CCKBCs beyond the hippocampal lesion may contribute to hyperexcitability and compromised network oscillations in TLE.

Introduction

Temporal lobe epilepsy (TLE) is the most common type of focal epilepsy and affects more than 60% of all epilepsy patients (Téllez-Zenteno and Hernández-Ronquillo, 2012). TLE is characterized by recurrent spontaneous seizures and associated comorbidities (Amlerova et al., 2013; Bui et al., 2018; Cánovas et al., 2011; Elger et al., 2004; Holmes, 2015; Kim et al., 2020). Although antiepileptic drugs (AEDs) are commonly used to control seizures, one third of epilepsy patients are refractory to AEDs (Kalilani et al., 2018). Temporal lobectomy is the most effective treatment for TLE when seizures are uncontrolled by AEDs, though there are risks and complications related to surgical therapy (Thom et al., 2010). The mechanisms underlying hyperexcitability, seizures, and impaired memory in TLE are not fully understood. Better understanding of such mechanisms will be critical for development of target molecules, cells, and circuits for therapeutic intervention in patients with refractory TLE.

Cholecystokinin-positive (CCK+) interneurons are a major group of GABAergic interneurons in the hippocampus that comprise the perisomatic CCK+ interneurons (i.e., CCK+ basket cells, CCKBCs) and dendritic CCK+ interneurons (e.g., Schaffer collateral-associated cells, SCA cells) (Bezaire and Soltesz, 2013; Lee et al., 2010). CCKBCs regulate excitability of hippocampal circuits and are critically involved in coordinated network oscillations in the hippocampus (e.g., theta rhythms), which support key functions of the hippocampus (e.g., episodic memory; Bezaire et al., 2016; Colgin, 2016; Freund and Katona, 2007). CCKBCs are vulnerable and CCK+ interneuron-mediated inhibition of principal cells is downregulated in animal models of TLE (Sun et al., 2014; Wyeth et al., 2010). Such changes in CCK+ interneuron circuits in the hippocampus may contribute to spontaneous seizures and memory deficits in TLE.

The unilateral intrahippocampal kainate (IHK) model of TLE is commonly used because it reproduces key features of human TLE: Spontaneous seizures controlled by AEDs, hippocampus-dependent memory deficits, and histopathological changes (e.g., hippocampal sclerosis; Rattka et al., 2013). Unlike animal models of TLE induced by systemic chemoconvulsant administration (e.g., pilocarpine or kainate) showing bilateral hippocampal sclerosis, the unilateral IHK model of TLE produces hippocampal sclerosis more restricted to the targeted ipsilateral hippocampal region, whereas the ipsilateral hippocampal region and contralateral hippocampus remote from the sclerotic region of the hippocampus appear to be relatively intact. Importantly, unilateral hippocampal sclerosis is observed in approximately 80% of patients with mesial temporal sclerosis (Alarcón and Valentín, 2010; Thom, 2014). Although most seizures arise from the sclerotic hippocampus in TLE patients with unilateral hippocampal sclerosis, the remaining seizures arise from other brain regions, including contralateral hippocampus (Bragin et al., 1999). Furthermore, reduced hippocampal theta network oscillations arise from ventral hippocampal circuits remote from the sclerotic region of the hippocampus in the dorsal unilateral IHK mouse model (Dugladze et al., 2007). These results collectively suggest that reorganization of neuronal circuits remote from hippocampal sclerosis may contribute to spontaneous seizures and compromised network oscillations in TLE. Indeed, a subset of hippocampal interneurons remote from the hippocampal lesion is vulnerable to injury in the IHK model (Marx et al., 2013), and neuronal degeneration in the contralateral dentate gyrus has been described (Groticke et al., 2008). However, it is unclear whether hippocampal CCK+ interneurons remote from the hippocampal lesion are vulnerable or whether surviving CCK+ interneurons manifest abnormal activity in the unilateral IHK mouse model.

In this study, we used immunostaining, confocal imaging, whole-cell patch-clamp recordings, video-EEG monitoring, and behavioral tests in a well-established unilateral IHK mouse model of TLE targeting dorsal hippocampus to test our hypothesis that ventral hippocampal CCK+ interneuron circuits remote from hippocampal sclerosis are compromised in the dorsal IHK mouse model. We focused on three main questions: 1) Is CCK+ interneuron number reduced in the ventral hippocampus in the dorsal IHK model; 2) do surviving CCKBCs in the IHK model manifest abnormal intrinsic and synaptic properties; and 3) does the IHK model result in downregulated CCK+ interneuron-mediated inhibition?

Section snippets

Animals

Young adult C57BL/6J mice (Jackson laboratory, Bar Harbor, ME, USA) were maintained at ambient temperature and humidity with a 14-h light/10-h dark cycle and fed standard chow ad libitum. Similar numbers of males and females were used for the study, thus reducing sex bias (Will et al., 2017). Thus, the combined data from both sexes are presented in this study. All animal procedures were approved by the Institutional Animal Care and Use Committees of the University of Kentucky and the University

IHK mouse model of TLE reproduced key properties of human TLE

The IHK mouse model of TLE reproduces key features of human TLE: Hippocampal sclerosis, behavioral and electrographic seizures, and cognitive dysfunction (Bui et al., 2018; Krook-Magnuson et al., 2013; Levesque and Avoli, 2013). In this study, we first determined whether our model reproduced previous findings. Behavioral seizures were observed after IHK-injected mice recovered from anesthesia. The mice initially displayed low levels of motor seizures such as repetitive mouth movement, head

Discussion

In this study, we found both structural and functional loss of CCK+ interneurons in ventral hippocampi in the dorsal IHK mouse model of TLE. The main findings in epileptic mice are: 1) CCK+ interneuron numbers were reduced in ventral hippocampi; 2) Axon terminals from CCKBCs targeting perisomatic regions of CA1 PCs were reduced in ventral hippocampi; 3) Surviving CCKBCs showed no major changes in their intrinsic and synaptic properties or connectivity with CA1 PCs; and 4) Carbachol-induced CCK+

Declaration of Competing Interest

None.

Acknowledgments

We thank H.E.S. Lewis and M.W. Young for technical assistance with the analysis of behavioral seizures, and Dr. M.B. Halmos for statistical advice. This work was supported by the College of Medicine, UAMS (startup funding to S.-H.L), Core Facilities of the Center for Translational Neuroscience at UAMS, Award P30 GM110702 from the IDeA program at NIGMS, and R01 NS092552 (to B.N·S). The sponsors had no role in study design, data collection, analysis and interpretation, or writing of this

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