The role of the cerebellum in drug-cue associative memory: functional interactions with the medial prefrontal cortex

Drug-induced Pavlovian memories are thought to be crucial for drug addiction because they guide behaviour towards environments with drug availability. Drug-related memory depends on persistent changes in dopamine-glutamate interactions in the medial pre- frontal cortex (mPFC), basolateral amygdala, nucleus accumbens core and hippocampus. Recent evidence from our laboratory indi- cated that the cerebellum is also a relevant node for drug-cue associations. In the present study, we tested the role that specific regions of the cerebellum and mPFC play in the acquisition of cocaine-induced preference conditioning. Quinolinic acid was used to manage a permanent deactivation of lobule VIII in the vermis prior to conditioning. Additionally, lidocaine was infused into the prelim- bic and infralimbic (IL) cortices for reversible deactivation before every training session. The present findings show, for the first time, that the cerebellum and mPFC might act together in order to acquire drug-cue Pavlovian associations. Either a dorsal lesion in lobule VIII or an IL deactivation encouraged cocaine-induced preference conditioning. Moreover, simultaneous IL-cerebellar deactivation prevented the effect of either of the separate deactivations. Therefore, similar to the IL cortex, neural activity in the cerebellum may be crucial for ensuring inhibitory control of the expression of cocaine-related memories

Cocaine-Induced Preference Conditioning: a Machine Vision Perspective

Existing work on drug-induced synaptic changes has shown that the expression of perineuronal nets (PNNs) at the cerebellar cortex can be regulated by cocaine-related memory. However, these studies on animals have mostly relied on limited manually-driven procedures, and lack some more rigorous statistical approaches and more automated techniques. In this work, established methods from computer vision and machine learning are considered to build stronger evidence of those previous findings. To that end, an image descriptor is designed to characterize PNNs images; unsupervised learning (clustering) is used to automatically find distinctive patterns of PNNs; and supervised learning (classification) is adopted for predicting the experiment group of the mice from their PNN images. Experts in neurobiology, who were not aware of the underlying computational procedures, were asked to describe the patterns emerging from the automatically found clusters, and their descriptions were found to align surprisingly well with the two types of PNN images revealed from previous studies, namely strong and weak PNNs. Furthermore, when the set of PNN images corresponding to every mice in the saline (control) group and the conditioned (experimental) group were characterized using a bag-of-words representation, and subject to supervised learning (saline vs conditioned mice), the high classification results suggest the ability of the proposed representation and procedures in recognizing these groups. Therefore, despite the limited size of the dataset (1,032 PNN images of 6 saline and 6 conditioned mice), the results support existing evidence on the drug-related brain plasticity, while providing higher objectivit

Involving the cerebellum in cocaine-induced memory: pattern of CFOS expression in mice trained to acquire conditioned preference for cocaine

Because of its primary role in drug-seeking, consumption and addictive behaviour, there is a growing interest in identifying the neural circuits and molecular mechanisms underlying the formation, maintenance and retrieval of drug-related memories. Human studies, which focused on neuronal systems that store and control drug-conditioned memories, have found cerebellar activations during the retrieval of drug-associated cue memory. However, at the pre-clinical level, almost no attention has been paid to a possible role of the cerebellum in drug-related memories. In the present study, we ought to fill this gap by aiming to investigate the pattern of neuronal activation (as revealed by cFos expression) in different regions of the prefrontal cortex and cerebellum of mice trained to develop conditioned preference for an olfactory stimulus (CS+) paired with cocaine. Our results indicate that CS+ preference was directly associated with cFos expression in cells at the apical region of the granule cell layer of the cerebellar vermis; this relationship being more prominent in some specific lobules. Conversely, cFos+ immunostaining in other cerebellar regions seems to be unrelated to CS+ preference but to other aspects of the conditioning procedure. At the prefrontal cortex, cFos expression seemed to be related to cocaine administration rather than to its ability to establish conditioned preference. The present results suggest that as it has been observed in some clinical studies, the cerebellum might be an important and largely overlooked part of the neural circuits involved in generating, maintaining and/or retrieving drug memories

Have we been ignoring the elephant in the room? Seven arguments for considering the cerebellum as part of addiction circuitry

Addiction involves alterations in multiple brain regions that are associated with functions such as memory, motivation and executive control. Indeed, it is now well accepted that addictive drugs produce long-lasting molecular and structural plasticity changes in corticostriatal-limbic loops. However, there are brain regions that might be relevant to addiction other than the prefrontal cortex, amygdala, hippocampus and basal ganglia. In addition to these circuits, a growing amount of data suggests the involvement of the cerebellum in many of the brain functions affected in addicts, though this region has been overlooked, traditionally, in the addiction field. Therefore, in the present review we provide seven arguments as to why we should consider the cerebellum in drug addiction. We present and discuss compelling evidence about the effects of drugs of abuse on cerebellar plasticity, the involvement of the cerebellum in drug-induced cue-related memories, and several findings showing that the instrumental memory and executive functions also recruit the cerebellar circuitry. In addition, a hypothetical model of the cerebellum's role relative to other areas within corticostriatal-limbic networks is also provided. Our goal is not to review animal and human studies exhaustively but to support the inclusion of cerebellar alterations as a part of the physiopathology of addiction disorder.This work was supported by grants and fellowships: FPU12/04059, PPF 2015 (15I082.01/1) and UJI (14I307.01/1). We also thank Timothy Attwood Gibbons for the English revision of the manuscript

Cocaine-induced plasticity in the cerebellum of sensitised mice

Rationale Prior research has accumulated a substantial amount of evidence on the ability of cocaine to produce short- and long-lasting molecular and structural plasticity in the corticostriatal-limbic circuitry. However, traditionally, the cerebellum has not been included in the addiction circuitry, even though growing evidence supports its involvement in the behavioural changes observed after repeated drug experiences. Objectives In the present study, we explored the ability of seven cocaine administrations to alter plasticity in the cerebellar vermis. Methods After six cocaine injections, one injection every 48 h, mice remained undisturbed for 1 month in their home cages. Following this withdrawal period, they received a new cocaine injection of a lower dose. Locomotion, behavioural stereotypes and several molecular and structural cerebellar parameters were evaluated. Results Cerebellar proBDNF and mature BDNF levels were both enhanced by cocaine. The high BDNF expression was associated with dendritic sprouting and increased terminal size in Purkinje neurons. Additionally, we found a reduction in extracellular matrix components that might facilitate the subsequent remodelling of Purkinje-nuclear neuron synapses. Conclusions Although speculative, it is possible that these cocaine-dependent cerebellar changes were incubated during withdrawal and manifested by the last drug injection. Importantly, the present findings indicate that cocaine is able to promote plasticity modifications in the cerebellum of sensitised animals similar to those in the basal ganglia.This work was supported by grants and fellowships: Ministerio de Economía y Competitividad [PSI2011- 29181], FPI-PREDOC2009/05, FPU12/04059, PPF 2013 (13I087.01/1) and UJI (P1.1B2011-42)

Cerebellar hallmarks of conditioned preference for cocaine

Pavlovian conditioning tunes the motivational drive of drug-associated stimuli, fostering the probability of those environmental stimuli to promote and trigger drug seeking and taking. Interestingly, different areas in the cerebellum are involved in the formation and long-lasting storage of Pavlovian emotional memory. Very recently, we have shown that conditioned preference for an odour associated with cocaine was directly correlated with cFOS expression in cells at the dorsal region of the granule cell layer of the cerebellar vermis. The main goal of the current investigation was to further extend the description of cFOS-IR patterns in cerebellar circuitry after training mice in a cocaine-odour Pavlovian conditioning procedure, including now the major inputs (the inferior olive and pontine nuclei) and one of the output nuclei (the medial deep nucleus) of the cerebellum. The results showed that the cerebellar hallmark of preference towards an odour cue associated to cocaine is an increase in cFOS expression in the dorsal part of the granule cell layer. cFOS-IR levels expressed in the granule cell layer of mice that did not show cocaine conditioned preference did not differ from the basal levels. Remarkably, mice subjected to a random cocaine-odour pairing procedure (the unpaired group) exhibited higher cFOS-IR in the inferior olive, the pontine nuclei and in the deep medial nucleus. Therefore, our findings suggest that inputs and the output of cerebellar circuitry are enhanced when contingency between the CS+ and cocaine is lacking

Cerebellar perineuronal nets in cocaine-induced pavlovian memory: Site matters

One of the key mechanisms for the stabilization of synaptic changes near the end of critical periods for experience-dependent plasticity is the formation of specific lattice extracellular matrix structures called perineuronal nets (PNNs). The formation of drug memories depends on local circuits in the cerebellum, but it is unclear to what extent it may also relate to changes in their PNN. Here, we investigated changes in the PNNs of the cerebellum following cocaine-induced preference conditioning. The formation of cocaine-related preference memories increased expression of PNN-related proteins surrounding Golgi inhibitory interneurons as well as that of cFos in granule cells at the apex of the cerebellar cortex. In contrast, the expression of PNNs surrounding projection neurons in the medial deep cerebellar nucleus (DCN) was reduced in all cocaine-treated groups, independently of whether animals expressed a preference for cocaine-related cues. Discriminant function analysis confirmed that stronger PNNs in Golgi neurons and higher cFos levels in granule cells of the apex might be considered as the cerebellar hallmarks of cocaine-induced preference conditioning. Blocking the output of cerebellar granule cells in a6Cre-Cacna1a mutant mice prevented re-acquisition, but not acquisition, of cocaine-induced preference conditioning. Interestingly, this impairment in consolidation was selectively accompanied by a reduction in the expression of PNN proteins around Golgi cells. Our data suggest that PNNs surrounding Golgi interneurons play a role in consolidating drug-related memories