Behavioral, imaging, and neurochemical correlates of the neonatal clomipramine model of depression in the rat

Abstract Depression is a widespread psychiatric disorder that represents a major cause of disability worldwide. Besides the significant mortality rate due to suicide, depression has been associated with an increased prevalence of coronary artery disease and type-2 diabetes. In spite of the accumulated knowledge on the heterogeneous collection of symptoms that characterize the disease, much has yet to be discovered about its pathophysiology. Risk factors include stressful life events, endocrine abnormalities, and genetic predispositions interacting with the environment. The development of animal models of depression have made it possible to study the behavioral and neurobiological correlates of the disease, although only a subset of the symptoms that characterize the human syndrome can be reproduced and studied in the animal. Clomipramine, a well-known tricyclic antidepressant, acts in an apparently paradoxical fashion when repeatedly administered to neonate rats. The treatment induces in the adult a series of behavioral and neurobiological effects similar to a depressive syndrome. The mechanisms underlying the phenomenon are still unknown, although hypotheses have been formulated on the potential depressogenic effects of clomipramine-dependent inhibition of REM sleep and alterations of the monoaminergic circuitry in a developing brain. In the present dissertation, I report the results of an in-depth investigation of the behavioral and neurobiological correlates of the neonatal clomipramine administration model of depression, not yet described or only partially analyzed in the literature. For each of the experiments, groups of Sprague-Dawley rats were subjected from the 5th to the 21st post-natal day to a protocol of clomipramine administration (20 mg/kg i.p., twice a day). The animals were compared with groups of rats treated with saline injections according to an identical schedule. Both groups of animals were tested for locomotor activity and despair behavior with the forced swim test, for anhedonic behavior with the sucrose preference test and for exploratory activity and anxiety trait with the elevated plus maze test. In all behavioral tests performed, clomipramine-treated animals showed abnormalities, although sometimes subtle, that can be attributed to depression-like symptoms, compared to saline-treated animals. Morphometric studies conducted by means of magnetic resonance imaging, performed to detect possible volumetric abnormalities in brain regions usually associated with depression, showed a significant decrease in brain volume in treated animals, together with a significant enlargement of the lateral ventricles, but no significant changes in the hippocampus. Investigations of the neurobiological correlates of the clomipramine treatment yielded results that include: 1) Brain-derived neurotrophic factor levels were significantly decreased in the hippocampus, but not in the cortex, of treated animals; 2) serotonin transporter expression, measured as optical density of the immunohistochemical labeling of the marker in brain sections, was decreased in the hippocampus and cingulate cortex of treated animals compared to the controls; 3) expression of the glial fibrillary acidic protein, a marker of astrocytes, was by neonatal treatment affected in relation with age and sex. On the other hand, hippocampal neurogenesis rate and hippocampal granule neuronal morphology were not significantly affected by treatment. The data suggest sex-dependent differences in the effects of clomipramine, with female rats being more responsive than males to the neonatal treatment. Furthermore, the behavioral symptoms as well as the neurobiological consequences of the treatment appeared to reduce with aging, to the point of almost complete recovery at an advanced age The neonatal clomipramine administration model was further characterized by testing the reaction of animals to an acute stressful event, in this case restraint stress realized by immobilizing animals with a wire mesh, on the rationale that such events can trigger or precipitate depression in humans and that a similar phenomenon may be observed in the animal model. For this study, a group of untreated rats, i.e. animals that were left undisturbed in their cages throughout their development, were added to the experimental design. An interaction between early life treatment and stress in adulthood was found in several experiments. Importantly, untreated animals showed substantial differences compared to the other two groups, suggesting that a significant fraction of the observed effects rather than being directly attributable to the pharmacological properties of clomipramine, may in fact depend on the procedure as a whole, including handling, injection discomfort, and separation from the mother. Overall, the studies reported here provide important elements for the evaluation of the efficacy and congruency of the early clomipramine treatment model of depression.