Research ArticleDynamics of Dendritic Spines in Dorsal Striatum after Retrieval of Moderate and Strong Inhibitory Avoidance Learning
Introduction
It has been shown that memory formation is intimately associated with spinogenesis. Indeed, numerous experiments have demonstrated that recent and remote memory are accompanied by changes in density and morphology of dendritic spines in various cerebral regions, such as the CA1 field and dentate gyrus of the hippocampus, the basolateral amygdala, and anterior cingulate cortex (e.g., Aceti et al., 2015, Eyre et al., 2003, Heinrichs et al., 2013, Leuner et al., 2003, Moser et al., 1994, O'Malley et al., 1998, O’Malley et al., 2000, Restivo et al., 2009). Of note is the paper by Hayashi-Takagi et al. (2015) where, using a synaptic optoprobe, they found labeling of recently potentiated spines in the primary motor cortex of rats after motor learning, and that shrinkage of the potentiated spines disrupted the learned response, thus giving strong support to the idea that spinogenesis represents a structural correlate of memory.
Although there is abundant literature showing that the dorsal striatum (DS) is a key element for consolidation of memory of a wide variety of learning tasks (for reviews see Divac and Öberg, 1979, Packard and Knowlton, 2002, Prado-Alcalá, 1995, White, 2009), the morphological substrates associated with these processes have been barely studied in this structure.
The work of Comery et al., 1995, Comery et al., 1996) showed that rearing rats in an enriched environment produced increased spine density in striatal medium spiny neurons (MSNs) of dorsolateral striatum (DLS), indicating that spinogenesis might occur in more structured types of learning. This is certainly the case for inhibitory avoidance (IA). We have shown that IA induces mushroom spinogenesis and reduction of thin spines in MSNs of DLS and dorsomedial (DMS) striatum in rats. These changes were evident at 6 h after training and persisted for at least 24 h afterward, coinciding with the initiation and maintenance of long-term memory (Bello-Medina et al., 2016). More recently, increased density of mushroom spines and reduced density of thin spines in dorsolateral striatal MSNs were also found when a response-training task in a plus maze was studied (Briones et al., 2018). By contrast, no changes in spine density were observed when rats were trained in a T-maze in a goal directed behavior and spatial learning, which involves DMS, and then switched to procedural or habitual performance, which involves DLS (Hawes et al., 2015). These results would indicate that the dynamics of striatal spinogenesis are dependent upon the type of learning task.
Other lines of research have demonstrated that strong training protects against treatments that commonly disrupt memory consolidation (Prado-Alcalá et al., 2012)1. Thus, post-training cholinergic blockade of the dorsal striatum (DS) produced by local infusions of atropine or scopolamine hindered consolidation of IA in rats, but this treatment did not interfere with memory when the animals had been trained with foot-shock of relatively high intensities (Cruz-Morales et al., 1992, Giordano and Prado-Alcalá, 1986). Furthermore, generalized inactivation of the DS, produced by post-training infusion of lidocaine, also interfered with memory consolidation of IA mediated by low-intensity foot-shock but not so after relatively high-intensity foot-shock (Pérez-Ruiz and Prado-Alcalá, 1989, Salado-Castillo, 2011).
There is evidence for striatal-dependent memory retrieval. Botreau et al. (2004) reported the occurrence of impaired processing of retrieval cues in a visual discrimination task after damage to the DS, and lesions of the caudate nucleus of rabbits deteriorates retrieval of trace eye blink conditioning (Flores and Disterhoft, 2013). Retrieval of memory is also protected by overtraining against amnestic treatments administered to the DS. Earlier findings of our group showed that cholinergic blockade and reversible inactivation of the caudate nucleus produced amnesia in cats that had achieved asymptotic performance of lever-pressing; overtraining, however, prevented this amnestic state (Prado-Alcalá and Cobos-Zapiaín, 1977, Prado-Alcalá and Cobos-Zapiaín, 1979). This protective effect was later generalized to rats trained with a high number of sessions in the lever-press task and then injected with scopolamine into the DS (Prado-Alcalá et al., 1980). Equivalent effects were reported by Kobayashi and Iwasaki (2000) who lesioned cholinergic striatal interneurons of rats that were trained or overtrained in the Morris water maze. By the same token, a significant impairment in allocentric tactile and spatial memory was produced by striatal lesions, but these impairments were significantly reduced with extensive preoperative training (Colombo et al., 1989).
The participation of several brain structures, such as the hippocampus, amygdala, entorhinal cortex, parietal cortex, and cingulate cortex, in retrieval of IA memory is well known (e.g., Izquierdo et al., 1997, Izquierdo et al., 2002), but little attention has been paid to the involvement of the striatum in retrieval of this aversively motivated learning task. It has been shown, for instance, that infusion of atropine into the DS before retrieval of IA disrupted the conditioned response (Prado-Alcalá et al., 1985), and retrieval of the same task increased the number of cholinergic M1 muscarinic receptors in this structure (Ortega et al., 1996). To further study this matter, we investigated if retrieval of inhibitory avoidance learning would induce changes in dendritic spine density and morphology in DMS and DLS, like those found after memory consolidation of this task (Bello-Medina et al., 2016); we reported that there was a direct relationship between the intensity of IA training and spine density. It has been shown that DLS activity is crucial for procedural learning and DMS is mainly involved in spatial/contextual learning (e.g., Devan and White, 1999, Packard and Goodman, 2012, Packard and Knowlton, 2002, Packard and McGaugh, 1996). In the present work, we predicted that these regions would show similar patterns of spine dynamics, for the reason that the inhibitory avoidance task entails both spatial and procedural components. Furthermore, we expected that stronger learning would induce greater changes in spine density and morphology than weaker learning. In our previous study (Bello-Medina et al., 2016) we also found significant changes in spine density and morphology in nucleus accumbens (NAc), but they were dependent on the aversive stimulation alone, and not on the associative process involved in IA. This result led us to investigate if this pattern of spine dynamics in the accumbens would also occur after retrieval of IA.
Section snippets
Experimental procedure
This study was carried out in accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health (National Research Council, 2011) and all experimental procedures were approved by the Animal Ethics Committee of Instituto de Neurobiología, Universidad Nacional Autónoma de México (Approval code: 098.A).
Training and retrieval
There were no significant differences in training latencies among the R-0, R-1, and R-3 groups (H(2) = 0.39, p = 0.82) (Fig. 1A). On the other hand, there were significant differences among the groups regarding escape latencies (H(2) = 19.84, p < 0.0001). The Dunn’s Multiple Comparison Test showed that R-0 had significantly higher escape latencies than R-1 and R-3 (p < 0.05 and p < 0.0001, respectively); the latter two groups did not differ from each other (Fig. 1B). These results were expected
Discussion
This is the first study describing changes in the density and morphology of dendritic spines in dorsal and ventral striatum induced by retrieval of IA learning.
When we investigated if in basal conditions there were different densities in the four striatal regions studied, we found that the quantification of spine density NAcC had a higher spine density than NAcS, DLS, and DMS. These results agree well with those of Meredith et al. (1992) who also found more spines in NAcC than in NAcS, and with
Conflict of interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Author contributions
PCB-M and RAP-A designed the experiments; PCB-M performed the experiments; GL-Q and RAP-A contributed reagents and materials; PCBM, ACM and RAP-A analyzed the data; PCB-M, ACM, CXR-L, MM-D, GL-Q and RAP-A wrote and revised the paper. All authors have approved the final version of the article.
Funding
The present work was supported by Dirección General de Asuntos del Personal Académico, Universidad Nacional Autónoma de México (IN203918) and Consejo Nacional de Ciencia y Tecnología (237570). This work was carried out in partial fulfillment of the requirements to obtain the Doctor’s Degree (Doctorado en Ciencias Biomédicas, Universidad Nacional Autónoma de México) by PCB-M, who was a recipient of a Graduate Scholarship from CONACYT (Becario 234103).
Acknowledgements
The authors thank Norma Serafín, Bernardino Osorio, Nydia Hernández, Nuri Aranda, Alberto Lara, Alejandra Castilla, Martín García, and María A. Carbajo for their excellent technical and administrative assistance, and Dr. M. Jeziorski for his helpful comments on the manuscript.
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