Neuronal circuits of passive and active fear behavior: focus on the role played by the medial prefrontal cortex in normal conditions and after antidepressant administration

• Grant number: 15/01544-0
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: June 01, 2015
• End date: May 31, 2016
• Field of knowledge: Biological Sciences - Pharmacology - Neuropsychopharmacology
• Principal Investigator: Hélio Zangrossi Júnior
• Grantee: Ana Beatriz Sant Ana Do Nascimento
• Supervisor: Cyril Herry
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Institution abroad: Institut National de la Santé et de la Recherche Médicale (Inserm), France
• Associated to the scholarship: 12/20016-6 - Involvement of 5-HT1A and 5-HT2C receptors of the dorsal hippocampus on the anxiolytic effect caused by antidepressant drugs, BP.DR

Abstract

Over the past years, numerous studies have identified a network of neuronal structures involved in the formation and expression of conditioned fear memories, detached the basolateral amygdala (BLA) and the hippocampus. In addition, the medial prefrontal cortex (mPFC) has also been enrolled in this neuronal circuit, through its dense interconnectivity with BLA and hippocampus. In the past decade, it has also become clear that distinct parts of the mPFC (mPFC prelimbic area -PL and mPFC infralimbic area- IL) differentially regulate the expression of freezing behavior. The present proposal aims at mapping the prefrontal circuitry and the mechanisms selectively involved in the control of freezing and active avoidance conditioned behaviors using a unique innovative cross-level approach combining cutting edge in vivo extracellular electrophysiological recording techniques, selective optogenetic manipulations and behavioral and pharmacological approaches. Our hypothesis is that specific prefrontal circuits (excitatory/inhibitory PL and IL neuronal circuits) differentially control freezing and active avoidance behavior. To test this hypothesis, we will use in vivo long-lasting single unit recordings simultaneously in PL and IL during the expression of these behaviors using a behavioral conditioned paradigm, that allows the assessment of freezing and active avoidance behaviors in the same animal. Subsequently, we plan to use optogenetic to identify and selectively manipulate specific excitatory/inhibitory prefrontal circuits controlling these fear behaviors. More specifically, aiming to determine the cell type of the recorded neurons, we will use a method that involves the expression of the light-activated opsins channelrhodopsin-2 (ChR2) or Archaeorhodopsins (ArchT) to restrict excitatory and inhibitory neuronal populations using viral vectors selected based on the neuronal type to be infected. Additionally, we will use mouse expressing Cre-recombinase, which are available at the host institute, under the specific promoters (for excitatory- CAMKII and for inhibitory neurons, the parvalbumin-PV and somatostatin- SOM). Using this system, ChR2 and ArchT expression only occur in cell expressing Cre and infected with the AAV, that is, depending on the Cre mouse line used, specifically in excitatory or inhibitory neurons. Beside the clear identification of cell types, using the above strategy and the data collected, we will activate/inhibit the cell body of excitatory/inhibitory neuronal populations at specific time-point during fear behavior in order to prevent or facilitate the development of freezing and active avoidance-evoked neuronal responses. Finally, to manipulate excitatory neurons targeting specific neuronal regions we will use a complementary strategy based on the co-injection of a conditional floxed AAV with Cre-dependent expression of ChR2 or ArchT in the input structure and a canine adeno-associated viral vector (CAV) that travels retrogradely through axons and expresses the Cre-recombinase (CAV-Cre) in the target region. With this strategy, only the neurons co-infected with the AAV and the CAV-Cre virus will express ChR2 and ArchT, and these are neurons projecting to a particular region. Once these neuronal identification and manipulation will be achieved, it will be then possible to evaluate the preferential target in the mPFC for the selective serotonin reuptake inhibitors (SSRI). To this end, we will perform simultaneous single unit recordings and acute or chronic SSRI systemic injections (fluoxetine) following the acquisition of our freezing/active avoidance behavioral paradigm, with the hope of contributing for the improvement of existing therapeutic approaches. (AU)