4. 4 Limitations
The study owns some limitations such as, firstly, the focus on ionotropic glutamate signaling and not, for instance, on the potential role of metabotropic mGluR5 receptors as suggested by EE-induced mGluR5 enhancement of hippocampal LTP (Buschler & Manahan-Vaughan, 2017). Therefore, ad-hoc molecular studies are needed on the entire glutamatergic components, as well as on other neurochemical pathways (e.g., BDNF, 5-HT) and brain regions (e.g., amygdala) relevant to rewarding and learning. Further, we did not investigate the mechanism underlying the expression change, whether due to changes in synthesis or degradation. We measured the glutamatergic changes only at 2 hours after the end of the brief EE exposure: we are aware that a series of timepoint assessments may provide a better understanding. We are also aware that only detecting protein expression cannot establish a causal relationship and that protein expression alone may not reflect functional mechanisms. Our rats were kept isolated thus raising the possibility that housing conditions may indeed influence the response of the glutamatergic synapse. For instance, we recently demonstrated in adult male mice that living environment (i.e., social vs isolation housing) influences the efficacy of EE/fluoxetine interaction in inhibiting relapse to sucrose (Pintori et al. , 2022b). Finally, we used only male rats in our study and, therefore, we do not know if our findings can be extended to female rats.