AAV-mediated hippocampal expression of short and long Homer 1 proteins differentially affect cognition and seizure activity in adult rats

Abstract

Homer proteins mediate molecular rearrangements leading to changes in spine morphology. This points to a role of Homer in learning and memory. Homer 1c features both the ligand binding domain and a coiled–coiled domain for self-multimerization. Homer 1a lacks the coiled–coiled domain. Here, we report a new isoform which we termed 1g, lacking the Homer ligand binding domain. We dissected the functional roles of the individual Homer 1 domains, encoded by Homer 1a, 1c, and 1g, in vivo. Recombinant adeno-associated virus (AAV)-mediated overexpression of these forms in the hippocampus of adult rats has opposing effects on learning behavior. Increased levels of Homer 1a impaired hippocampal-dependent memory, while Homer 1g and 1c slightly enhanced memory performance. Homer 1g induced anxiety. Moreover, AAV-Homer 1a animals showed attenuation of electrographic seizures in a model of status epilepticus. These results suggest that Homer 1 proteins play an active role in behavioral plasticity.

Introduction

Long-lasting forms of memory are thought to be mediated by modifications in synapses that are induced by particular patterns of activity and involve a molecular cascade consisting of receptor activation, new gene expression, and protein synthesis (Steward and Worley, 2002). The molecular mechanisms underlying such long-lasting synaptic modifications remain to be fully defined.

The Homer family of proteins are a major component of the postsynaptic density (PSD) of excitatory synapses in the central nervous system. The products of three Homer genes (Homer 1–3) are dendritically enriched scaffold proteins, with a proposed role in the regulation of synaptic excitation and as such are implicated in the molecular rearrangement underlying synaptic plasticity (Fagni et al., 2002).

All Homer proteins feature a common N-terminal Enabled/Vasp homology (EVH1) domain, which binds a proline-rich motif in inositol 1,4,5-trisphosphate receptors (IP₃Rs), ryanodine receptors, type-1 metabotropic glutamate receptors (mGluRs) (Feng et al., 2002, Tu et al., 1998), and in Shank proteins that are scaffolding components of the NMDA receptor-associated PSD-95 complex (Naisbitt et al., 1999). Long forms additionally encode a C-terminal coiled–coiled (CC) domain followed by leucine zipper motifs facilitating oligomerization (Kato et al., 1998, Xiao et al., 1998). By self-aggregation and EVH1-mediated binding of their ligands, long Homer proteins physically link plasma membrane channels with intracellular Ca²⁺-stores thereby modulating excitatory signaling (Tu et al., 1998, Yuan et al., 2003). Expression of the immediate early gene (IEG) product Homer 1a is induced by both physiological and excessive neuronal activity (Brakeman et al., 1997). Homer 1a lacks the CC domain indicating it can compete with the long Homer proteins for binding to signaling components and thus functions as a natural dominant-negative regulator of postsynaptic Ca²⁺-dynamics (Fagni et al., 2002). Recent work revealed that Homers do not function as simple scaffolds, as deletion of Homer 2 or 3 does not disrupt polarized localization of IP₃Rs and other Ca²⁺-signaling proteins in peripheral cells but affects the efficiency of signal transduction (Shin et al., 2003). Moreover, deletion of Homer 1 increases spontaneous Ca²⁺-influx mediated by disturbed gating of TRPC1 ion channels, which are widely expressed in brain (Yuan et al., 2003). Recently, the product of the X-linked mental retardation (MRX) gene, oligonephrin-1, has been shown to affect dendritic spine morphogenesis and to interact with Homer 1 at the PSD (Govek et al., 2004).

However, the functional significance of Homer proteins with respect to behavioral plasticity is unclear and few studies have investigated the role of Homer proteins in vivo using transgenic approaches. Deletion of the single Homer gene in Drosophila impairs behavioral plasticity (Diagana et al., 2002) and mosaic overexpression of Homer 1a in the CNS of transgenic mice moderately retards epileptogenesis in the kindling model of epilepsy (Potschka et al., 2002). Deletion of Homer 1 or Homer 2 in mice cause the same increase in sensitivity to cocaine-induced locomotion, conditioned reward, and augmented glutamate transmission in nucleus accumbens as that elicited by withdrawal from repeated cocaine administration. Moreover, adeno-associated virus (AAV)-mediated restoration of Homer 2 in the accumbens of Homer 2 KO mice reversed the cocaine-sensitized phenotype (Szumlinski et al., 2004).

On the basis of this, we hypothesized that overexpression of Homer 1 proteins in the hippocampus, a brain region centrally involved in learning and epilepsy, would impact on behavioral plasticity. To address this, we have used an AAV-based gene transfer approach to overexpress Homer 1a, 1c, or a novel isoform, Homer 1g, in the hippocampus of adult rats. Here, we show that overexpression of Homer 1a leads to learning deficits but prevents status epilepticus (SE), while Homer 1c and 1g enhance cognitive performance but do little to alter susceptibility to SE.