Layer-specific modulation of entorhinal cortical excitability by presubiculum in a rat model of temporal lobe epilepsy

Abstract

Abbasi S, Kumar SS. Layer-specific modulation of entorhinal cortical excitability by presubiculum in a rat model of temporal lobe epilepsy. J Neurophysiol 114: 2854 -2866, 2015. First published September 16, 2015 doi:10.1152/jn.00823.2015 is the most common form of epilepsy in adults and is often refractory to antiepileptic medications. The medial entorhinal area (MEA) is affected in TLE but mechanisms underlying hyperexcitability of MEA neurons require further elucidation. Previous studies suggest that inputs from the presubiculum (PrS) contribute to MEA pathophysiology. We assessed electrophysiologically how PrS influences MEA excitability using the rat pilocarpine model of TLE. PrS-MEA connectivity was confirmed by electrically stimulating PrS afferents while recording from neurons within superficial layers of MEA. Assessment of alterations in PrS-mediated synaptic drive to MEA neurons was made following focal application of either glutamate or NBQX to the PrS in control and epileptic animals. Here, we report that monosynaptic inputs to MEA from PrS neurons are conserved in epileptic rats, and that PrS modulation of MEA excitability is layer-specific. PrS contributes more to synaptic inhibition of LII stellate cells than excitation. Under epileptic conditions, stellate cell inhibition is significantly reduced while excitatory synaptic drive is maintained at levels similar to control. PrS contributes to both synaptic excitation and inhibition of LIII pyramidal cells in control animals. Under epileptic conditions, overall excitatory synaptic drive to these neurons is enhanced while inhibitory synaptic drive is maintained at control levels. Additionally, neither glutamate nor NBQX applied focally to PrS now affected EPSC and IPSC frequency of LIII pyramidal neurons. These layer-specific changes in PrS-MEA interactions are unexpected and of significance in unraveling pathophysiological mechanisms underlying TLE. temporal lobe epilepsy; presubiculum; glutamate application; hyperexcitability; electrophysiology TEMPORAL LOBE EPILEPSY (TLE) is characterized by spontaneous recurrent seizures emanating from temporal lobe foci. It is the most common form of epilepsy in adults, and is often associated with drug-resistant seizures (Engel 1998;Engel et al. 2008).

Functional Connectivity of the Parasubiculum and Its Role in Temporal Lobe Epilepsy

“…The cut dorsal surface of the brain was adhered to the cutting platform using cyanoacrylate glue. This cutting angle was chosen to increase the likelihood of preserving PrS/Par projections to MEA in acute brain slices (Abbasi and Kumar, 2014;Abbasi and Kumar, 2015;Canto et al, 2012;Honda and Ishizuka, 2004;Tolner et al, 2007). Semi-horizontal slices (400 μm) were prepared using a microslicer (VT1000S, Leica, Germany) in the chilled cutting solution and allowed to equilibrate in oxygenated artificial cerebrospinal fluid (aCSF) containing (in mM) 126 NaCl, 26 NaHCO 3 , 3 KCl, 1.25 NaH 2 PO 4 , 2 MgSO 4 , 2 CaCl 2 , 0.25 L-Glutamine, and 10 D-glucose (pH 7.4), first at 32°C for 1 h and subsequently at room temperature before being transferred to the recording chamber.…”

“…Experimental compounds were focally applied within the Par / PrS using CESOP (Abbasi and Kumar, 2015) to maximize neuronal accessibility while minimizing spillover into neighboring nuclei, especially under turbulent or non-laminar flow conditions in the recording chamber. This system consisted of two perfusion pipettes piggy-backed onto one another and staggered such that the longer, narrower pipette (~250 μm) served as the inflow line, while the other, with a slightly larger tip diameter (~750 μm), served as the outflow.…”

“…CESOP enables focal application of pharmacological compounds to a brain region in submerged slices, while restricting their spillover to juxtaposed regions ( Fig. 3B (Abbasi and Kumar, 2015), 3D1-3D3). Because ambient network activity could vary between slices, glutamate (100 μM) application was followed by NBQX (10 μM) using CESOP, the pharmacological equivalent of a pulse function, to maximally enhance and then suppress overall synaptic activity of neurons in one region while recording from another neighboring region of interest (Fig.…”

“…Recently, we classified PrS neurons into distinct cell types based on electrophysiological criteria, establishing baselines for intrinsic properties mediating action potential waveforms and synaptic drive in each of the neuron types (Abbasi and Kumar, 2013). We assessed the physiological state of PrS neurons in animals with TLE directly to determine which of these neuronal populations are affected along with the nature of their physiological alterations (Abbasi and Kumar, 2014;Abbasi and Kumar, 2015). Through the present study, we showcase the Par by assessing a) the diversity of its neurons electrophysiologically; b) their intrinsic excitability as well as their excitatory and inhibitory synaptic drive under normal and epileptic conditions and c) their connectivity and influence over the dominant cell types in the PrS (LII and LIII) and the MEA (LII and LIII) using CESOP, a novel microfluidic technique for focal drug application (Abbasi and Kumar, 2015).…”

“…We assessed the physiological state of PrS neurons in animals with TLE directly to determine which of these neuronal populations are affected along with the nature of their physiological alterations (Abbasi and Kumar, 2014;Abbasi and Kumar, 2015). Through the present study, we showcase the Par by assessing a) the diversity of its neurons electrophysiologically; b) their intrinsic excitability as well as their excitatory and inhibitory synaptic drive under normal and epileptic conditions and c) their connectivity and influence over the dominant cell types in the PrS (LII and LIII) and the MEA (LII and LIII) using CESOP, a novel microfluidic technique for focal drug application (Abbasi and Kumar, 2015). To gain insights into how Par neuronal function and circuitry might be altered in TLE, we reassessed the physiological state of Par neurons and their connections in epileptic animals using the well-established pilocarpine model of TLE (Buckmaster, 2004).…”