*3.4. Limitations of the Study and Future Perspectives*

We evaluated changes of behavioral responses and mRNA expression after a short period (1 h) of drug exposure. This approach may be questionable, however there are many studies that support our observations. For instance, the mRNA expression of metabotropic glutamate receptors was found to be upregulated 1 h after treatment in mice DRG neurons [106]. Ambalavanar et al. [107] were able to detect changes in CGRP mRNA levels in rat's TG even 30 minutes after complete Freund's adjuvant injection. Furthermore, Nesic and co-workers [108] reported change in the mRNA signal of cytokines 1 h after treatment with MK-801, a NMDA receptor antagonist, in the spinal cord of rats subjected to spinal cord injury.

Nevertheless, to elucidate and confirm the present findings, additional experiments with different techniques are necessary. It will be interesting to evaluate in this model the effects of ADM\_12 at later time points, as well as after chronic treatment. Another limitation of the present study is the absence of a time course of the expression of CRGP and SP. This was motivated by the ethical and organizational need to keep the number of rats as low as possible. However, based on the present findings, it seems important to address in future studies the parallel evaluation of mRNA and protein expression of CGRP and SP in order to elucidate more clearly the role of these neuropeptides in peripheral and central sites.
