The T-type calcium channel antagonist Z944 rescues impairments in crossmodal and visual recognition memory in Genetic Absence Epilepsy Rats from Strasbourg
Introduction
Childhood absence epilepsy (CAE) accounts for approximately 8% of all epilepsies in school-aged children and is characterized by a disruption in consciousness sometimes with mild clonic movements and automatisms (Pavone et al., 2001). Although CAE was originally considered a benign disorder (Dieterich et al., 1985), recent research has shown that children with absence epilepsy have co-morbid behavioral (Caplan et al., 2009) and cognitive (Caplan et al., 2009, Henkin et al., 2005, Killory et al., 2011, Loughman et al., 2014, Mandelbaum and Burack, 1997, Pavone et al., 2001) symptoms. In particular, visual skill and visual memory deficits are consistently observed in CAE (Nolan et al., 2004, Pavone et al., 2001, Siren et al., 2007). Genetic Absence Epilepsy Rats from Strasbourg (GAERS) is an animal model closely resembling CAE. GAERS not only demonstrate recurrent non-convulsive seizures with bilateral and synchronous spike-and-wave discharges (SWD) characteristic of CAE (Marescaux et al., 1992) but also the anxiety and psychiatric-like phenotypes associated with epilepsy (Bouilleret et al., 2009, Dezsi et al., 2013, Jones et al., 2008, Jones et al., 2010, Marks et al., 2016; Powell et al., 2014; but see also Marques-Carneiro et al., 2014). Similar to CAE, GAERS exhibit altered cognition with increased performance observed in fear conditioning and two-way active avoidance tests (Getova et al., 1997, Marks et al., 2016), performance deficits observed for latent inhibition and extinction of conditioned fear (Marks et al., 2016), and delayed acquisition of spatial reference and working memory in a Morris water maze (Marques-Carneiro et al., 2016). Although progress has been made in characterizing cognitive alterations in GAERS, previous research has observed behavioral alterations using aversive Pavlovian and operant conditioning or stress-inducing tasks. Further, the mechanisms mediating alterations in cognition and behavior in CAE models are poorly understood.
Spontaneous object recognition tasks rely on the natural curiosity of rodents to explore novelty and are advantageous in that they measure learning and memory without prior training and are non-aversive (Cazakoff et al., 2010, Dere et al., 2007, Winters et al., 2008). Crossmodal object recognition (CMOR) is a recently developed memory task that examines not only purely visual and tactile memory, but also requires rodents to integrate sensory information about objects gained from tactile exploration to enable visual recognition (Ballendine et al., 2015, Winters and Reid, 2010, Reid et al., 2014). Both visual learning and memory deficits and sensory integration deficits are observed in CAE and psychiatric disorders that are highly comorbid with epilepsy (Heinrichs and Zakzanis, 1998, Stone et al., 2011, Williams et al., 2010, Wood et al., 2002). Given the specific learning and memory deficits observed in humans with epilepsy, examining CMOR performance in GAERS will broaden our understanding of the cognitive alterations associated with this absence epilepsy model.
In GAERS, a gain-of-function missense mutation in the Cav3.2 T-type calcium channel gene contributes to seizure activity (Powell et al., 2009). Further, administration of the pan-T-type calcium channel blocker, Z944, attenuates absence seizures in GAERS (Tringham et al., 2012). Previous research has shown that Cav3.2 T-type calcium channel deficient mice are impaired on novel and spatial object recognition tasks although working memory was unaffected (Gangarossa et al., 2014). Lesions of the perirhinal cortex (PRh) and posterior parietal cortex also disrupt CMOR with the PRh particularly relevant to visual recognition memory (Winters and Reid, 2010). Given that T-type calcium channels are expressed throughout the cerebral cortex with particularly dense expression in the PRh (Talley et al., 1999), investigating the role of T-type channels in mediating crossmodal recognition memory performance is warranted. We have recently demonstrated that Z944 administration has a profound effect on prepulse inhibition, a cognitive test that measures sensorimotor gating, in GAERS, NEC, and Wistar rats (Marks et al., in press). Taking into consideration the robust effects of Z944 on behavior, the objective of the present study was to examine the dose-dependent effect of Z944 on CMOR performance in GAERS and NEC rats. Experiments confirmed a deficit in both visual and crossmodal memory performance in GAERS compared to NECs. As anticipated, visual and crossmodal memory performance in GAERS was rescued through blockade of T-type calcium channels by Z944.
Section snippets
Animals
Male and female GAERS and NEC (University of Saskatchewan Lab Animal Services Unit, Saskatoon, Canada) (Marks et al., 2016) were used. All rats were group housed (2 or 3 per cage) in standard polypropylene cages in a temperature controlled (21 °C) colony room on a 12/12 h light/dark cycle. Experimental procedures were carried out during the light phase (lights on at 07:00 h), and food (Purina Rat Chow) and water were available ad libitum. This work was approved by the University of Saskatchewan's
Characterization of absence seizures during crossmodal testing
GAERS and NECs were first assessed for SWDs with freely moving EEG similar to previous studies (Marks et al., 2016, Powell et al., 2014). During a 20 min habituation period in the Y-shaped box prior to the crossmodal sample phase, GAERS displayed spontaneous SWDs resembling those described previously (Marescaux et al., 1992, Marks et al., 2016, Powell et al., 2009, Powell et al., 2014). NECs did not display absence seizures or SWD activity (Fig. 2). SWD were observed during 15.2 ± 2.3% of the
Discussion
In a series of experiments, we characterized the effects of acute systemic treatment with the T-type calcium channel blocker, Z944, on CMOR memory performance in GAERS and NEC rats. Recognition memory deficits in the tactile, visual and CMOR tests were observed in drug naïve GAERS relative to NECs, although the deficit in tactile recognition memory was less pronounced (Fig. 3, Fig. 4). In GAERS, Z944 had a robust dose-dependent effect on crossmodal test performance with a significant reversal
Conclusion
In a series of experiments, we demonstrated visual and CMOR memory deficits in drug naïve GAERS relative to NECs. Z944 rescued visual and CMOR deficits in GAERS, whereas, NEC animals showed impaired visual and CMOR performance in response to Z944 treatment. Tactile recognition memory was moderately increased by Z944 treatment in both strains. In general, object recognition memory specifically dependent on visual processing was robustly affected by Z944 treatment in both the GAERS and NEC
Funding and disclosure
This research was supported by a Brain Canada Multi-Investigator Research Initiative Grant with matching support from Genome British Columbia, the Michael Smith Foundation for Health Research, and the Koerner Foundation (PI: TPS). Work in the laboratory of TPS is supported by an operating grant from the Canadian Institutes for Health Research (CIHR; #10677) and a Canada Research Chair in Biotechnology and Genomics-Neurobiology. Work in the laboratory of JGH is supported by an operating grant
Acknowledgements
We wish to thank Mr. Jeff LeDue for assistance in developing EEG seizure analysis software and Mr. Brendan G. Murray and Ms. Nadine Zabder for assistance with video scoring.
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