4.2 Effects of brief EE exposure in the Nucleus accumbens
Brief EE exposure caused post-synaptic glutamatergic signaling changes in the NAc that are opposite compared to mPFC. In particular, EE induced an overall reduction of NMDARs subunits, both accessory (GluN2A, GluN2B) and obligatory (GluN1), in the PSD. Notably, no changes were observed in the expression of these subunits in the whole homogenate, suggesting that receptor synthesis is not affected while receptor retention is reduced as a consequence of brief EE exposure. This possibility is corroborated by the fact that the expression of SAP102, the scaffolding protein specific for NMDA receptors, is not increased to compensate for reduced expression, thus leading to unstable synapses. These results suggest that NMDA-mediated neurotransmission is depotentiated in the NAc, evidence also strengthened by reduced expression of PSD95 that, besides being a scaffolding protein, is an integral protein of the glutamate synapse centrally involved in multiple aspects of synaptic function (Vallejo et al. , 2017). Moreover, since structural changes of dendritic spines are sustained by PSD95 and Arc/Arg3.1 (Newpher et al. , 2018), their reduced expression suggests that a brief EE exposure might have reduced the density of dendritic spines contributing to the toning down of the NMDA receptors-mediated glutamate neurotransmission.
A different pattern can be observed for AMPA receptors. We found a significant reduction only in the expression of the GluA2 subunit, thus leading to an increase in GluA1/GluA2 ratio. This enhancement of GluA2-lacking AMPA receptors reflects a higher Ca2+permeability AMPA-mediated in the excitatory synapse, representing a metaplastic phenomenon, as it is established that the up-regulation of calcium-permeable AMPARs (CP-AMPAR) is able to change the threshold for different forms of plasticity (Liu & Zukin, 2007). While CP-AMPARs are expressed at low levels under basal conditions, their expression in the excitatory neurons increased in response to neuronal activity and neuronal insults. Importantly, CP-AMPARs in the NAc are involved in drug-seeking behaviors (see review (Neuhofer & Kalivas, 2018)). For instance, it has been demonstrated that the increase of NAc CP-AMPARs plays a critical role in the incubation of cocaine craving (Conradet al. , 2008; Wolf & Ferrario, 2010; Lee et al. , 2013; Caffino et al. , 2021). These data show that even a brief EE exposure may influence, through changes in accumbal glutamate homeostasis, salience attribution, and behavioral response to reward-associated stimuli (i.e., cue, context), ultimately driving maladaptive reward-related behaviors. In line with this evidence, the enhancement of CP-AMPARs levels observed in the NAc after brief EE exposure is consistent with the potentiation of Cx-induced sucrose-seeking previously reported (Pintori et al. , 2022a). Of note, the excessive calcium influx via EE-induced increase of CP-AMPARs can contribute to NMDA receptor inactivation, as previously demonstrated (Legendre et al. , 1993; Paoletti et al. , 2013; Sibarov & Antonov, 2018), thus further reducing the postsynaptic response.
We also found no changes in glutamate release as shown by the lack of effect on vGluT1. In addition, EE exposure leads to increased GLT-1 levels presumably to limit glutamate spillover and to prevent activation of extra-synaptic glutamate receptors that can impair synaptic plasticity (Scimemi et al. , 2009).