Dorsal striatum and the temporal expectancy of an aversive event in Pavlovian odor fear learning

Highlights

• Respiratory frequency is a good index of temporal expectancy of an aversive event in odor fear learning.

• Dorsal striatum inactivation modulates adaptation of the respiration temporal pattern to a new odor-shock time interval.

• Dorsal striatum dopamine levels decrease when a new interval duration is presented.

• Using a non-motor task allowed us to uncover a novel role for the striatum in interval timing.

Abstract

Interval timing, the ability to encode and retrieve the memory of intervals from seconds to minutes, guides fundamental animal behaviors across the phylogenetic tree. In Pavlovian fear conditioning, an initially neutral stimulus (conditioned stimulus, CS) predicts the arrival of an aversive unconditioned stimulus (US, generally a mild foot-shock) at a fixed time interval. Although some studies showed that temporal relations between CS and US events are learned from the outset of conditioning, the question of the memory of time and its underlying neural network in fear conditioning is still poorly understood. The aim of the present study was to investigate the role of the dorsal striatum in timing intervals in odor fear conditioning in male rats. To assess the animal’s interval timing ability in this paradigm, we used the respiratory frequency. This enabled us to detect the emergence of temporal patterns related to the odor-shock time interval from the early stage of learning, confirming that rats are able to encode the odor-shock time interval after few training trials. We carried out reversible inactivation of the dorsal striatum before the acquisition session and before a shift in the learned time interval, and measured the effects of this treatment on the temporal pattern of the respiratory rate. In addition, using intracerebral microdialysis, we monitored extracellular dopamine level in the dorsal striatum throughout odor-shock conditioning and in response to a shift of the odor-shock time interval. Contrary to our initial predictions based on the existing literature on interval timing, we found evidence suggesting that transient inactivation of the dorsal striatum may favor a more precocious buildup of the respiratory frequency’s temporal pattern during the odor-shock interval in a manner that reflected the duration of the interval. Our data further suggest that the conditioning and the learning of a novel time interval were associated with a decrease in dopamine level in the dorsal striatum, but not in the nucleus accumbens. These findings prompt a reassessment of the role of the striatum and striatal dopamine in interval timing, at least when considering Pavlovian aversive conditioning.

Introduction

Learning time intervals is crucial to survival and goal reaching across the phylogenetic tree. In Pavlovian fear conditioning, an initially neutral stimulus predicts the arrival of an aversive unconditioned stimulus, after a time interval that is encoded, a process pertaining to interval timing (Molet and Miller, 2014, Kirkpatrick and Balsam, 2016). How the brain processes and encodes such information remains poorly understood (Merchant et al., 2013, Tallot and Doyère, 2020). From a neurobiological point of view, there is substantial support for the involvement of the dorsal striatum and its dopaminergic inputs in interval timing (Buhusi & Meck, 2005), both from lesion experiments of the dorsal striatum or intrastriatal infusion of dopaminergic antagonists (De Corte et al., 2019, Meck, 2006), and from lesion or optogenetic manipulation of the substantia nigra pars compacta (SNc), the primary source of dorsostriatal dopamine (Meck, 2006, Soares et al., 2016). Electrophysiological recordings of single neurons in the dorsal striatum of rats also show that striatal neurons firing rate is correlated with the duration of the time interval between signaling cue and reward (Bakhurin et al., 2017, Gouvêa et al., 2015, Matell et al., 2003, Mello et al., 2015), and intrastriatal muscimol infusions produce an impairment in the animals’ ability to discriminate durations (Gouvêa et al., 2015). In the context of fear conditioning, using 2-Deoxyglucose (2-DG) metabolic mapping we previously showed that odor-shock pairing in rats was associated with an increase in 2-DG uptake in the dorsal striatum (Boulanger Bertolus et al., 2014). Furthermore, recent work recording oscillatory neural activity in the dorsal striatum in Pavlovian fear conditioning correlated its maximum power in theta and gamma bands with the time at which the rat expected the aversive stimulus (Dallérac et al., 2017). Timing in fear conditioning is also associated with plasticity in the striatum (Dallérac et al., 2017).

Notably, a majority of studies investigating the neurobiological substrate of interval timing in rodent models rely on operant conditioning. Such protocol relies heavily on the motor response of the subject, which could bias our understanding of the neural substrate of interval timing per se. Indeed, the striatum and cortico-striatal inputs are also a neural substrate for motor and procedural learning (Barnes et al., 2005, Koralek et al., 2013, Martiros et al., 2018), and action selection when the task involves temporal discrimination (Howard, Li, Geddes, & Jin, 2017). To avoid these possibly confounding factors, we used a non-striatum-dependent behavioral measure, the respiratory frequency, to assess the animal’s interval timing ability in a Pavlovian fear conditioning associating an odor to a mild footshock. Indeed, the pattern of respiratory frequency has been shown to be a good index of the animal’s temporal expectation of the shock arrival (Boulanger Bertolus et al., 2014, Dupin et al., 2020, Shionoya et al., 2013). This allowed us to look more closely at the role of dorsal striatum and its dopamine level in the initial acquisition of an interval duration, as well as when a change in this duration is applied. Based on the existing literature, we made two hypotheses 1) inactivating the dorsal striatum should impair timing behavior and its adaptation to a new interval duration and 2) dopamine level in the dorsal striatum should increase when a new interval duration is introduced. We report that, while a transient inactivation of the striatum did alter the expression of the temporal pattern of respiration, and dopamine level in the striatum was indeed modulated during the learning of a new duration, the directions of the effects were the opposite of those hypothesized. These findings prompt a reconsideration of the role of the striatum and striatal dopamine in interval timing, at least when considering Pavlovian aversive conditioning.