**1. Introduction**

Cannabidiol (CBD) is one of the most abundant phytocannabinoids present in the Cannabis sativa plant, devoid of psychotomimetic effects [1]. In addition, CBD antagonizes several of the psychoactive effects of the Δ9-tetrahydrocannabinol (Δ9-THC), the major psychoactive compound of Cannabis sativa [2,3]. CBD exhibits a broad-spectrum pharmacological profile that makes it a potential treatment for several psychiatric and neurodegenerative disorders [4,5]. In fact, while CBD displays low affinity for cannabinoid CB1 and CB2 receptors [6], it can act as a negative allosteric modulator at these receptors [7,8]. In addition, its mechanism of action involves, at least, inhibition of anandamide hydrolysis [9], as well as action at the vanilloid receptor 1 (TRPV1) [9], the serotoninergic

**Citation:** Mottarlini, F.; Fumagalli, M.; Castillo-Díaz, F.; Piazza, S.; Targa,G.; Sangiovanni, E.; Pacchetti, B.; Sodergren, M.H.; Dell'Agli, M.; Fumagalli, F.; et al. Single and Repeated Exposure to Cannabidiol Differently Modulate BDNF Expression and Signaling in the Cortico-Striatal Brain Network. *Biomedicines* **2022**, *10*, 1853. https://doi.org/10.3390/ biomedicines10081853

Academic Editor: Wesley M. Raup-Konsavage

Received: 7 July 2022 Accepted: 29 July 2022 Published: 1 August 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

5-HT1A receptor [10], and PPARγ receptors [11]. Friedman et al., reviewed the pharmacology of cannabinoids in neurological disorders, underlining the criticisms related to the bioavailability of these highly lipophilic molecules, such as the way of administration. With regards to CBD, the authors concluded that the bioavailability of oral administration in humans is poor (6–19%) and variable, according to several clinical studies [12,13]. For this reason, many in vivo studies concerning the effects of CBD in the central nervous system (CNS), including ours, considered other ways of administration, such as intraperitoneal (i.p.) injection. Regardless of the mode of administration, a clear characterization of the brain distribution of CBD and of its neuroplastic effects is still lacking.

Brain-derived neurotrophic factor (BDNF) belongs to a group of proteins termed neurotrophins, which play a pleiotropic role in the CNS. Besides its role during neurodevelopment and its neuroprotective properties, BDNF and its high affinity receptor TrkB are crucial regulators of neuroplasticity [14]. BDNF-induced activation of downstream signaling cascades regulates different cellular processes including neuroprotection, cognition, and stress coping [15]. Furthermore, modulation of BDNF levels may be critical for treatment of both psychiatric [16] and neurodegenerative disorders [17].

The organization of the BDNF gene is complex, exhibiting several 5 non-coding exons, each with a separate promoter region that triggers the transcription of a common 3 exon and encoding for the same protein [18]. The functions of these multiple variants have not ye<sup>t</sup> been fully clarified but, indeed, specific transcripts may undergo different intracellular targeting. Thus, it appears that BDNF function is controlled in a dynamic manner not only at the transcriptional and translational level, but also via specific mRNA targeting as well as processing and secretion of its protein [19,20]. In fact, the cleavage of the BDNF precursor gives rise to the mature form of the neurotrophin (mBDNF, 14 kDa), which is anterogradely transported to its target neurons [21,22]. This is an important point in the overall regulation of the BDNF system as, for instance, BDNF is not synthesized in the striatum. After its release, BDNF binds to its high-affinity receptor TrkB that, after autophosphorylation, stimulates downstream pathways, primarily phosphoinositide 3-kinase (PI3-K) and mitogen-activated protein kinase (MAPK) pathways. Several botanicals and natural compounds, including CBD, have been shown to modulate neuroplasticity acting on BDNF levels [23] in animal models of several disorders such as depression [24], schizophrenia [25], addiction [26], ischemia [27], and Alzheimer's disease [28]. However, information on the fine-tuned regulation of BDNF in the action of CBD is still fragmentary.

Accordingly, we performed a dose−response study in rats to evaluate the rapid modulation (1 h) of the neurotrophin as well as its downstream signaling following a single injection of CBD. In addition, we exposed rats to repeated CBD treatment (five consecutive injections, once a day) with sacrifice 24 h after the last injection, to investigate a potential drug-free time point, focusing our attention on CBD-induced modulation of BDNF in the cortico-striatal pathway.

#### **2. Materials and Methods**
