Memory formation and retrieval of neuronal silencing in the auditory cortex

Abstract

Sensory stimuli not only activate specific populations of cortical neurons but can also silence other populations. However, it remains unclear whether neuronal silencing per se leads to memory formation and behavioral expression. Here we show that mice can report optogenetic inactivation of auditory neuron ensembles by exhibiting fear responses or seeking a reward. Mice receiving pairings of footshock and silencing of a neuronal ensemble exhibited a fear response selectively to the subsequent silencing of the same ensemble. The valence of the neuronal silencing was preserved for at least 30 d and was susceptible to extinction training. When we silenced an ensemble in one side of auditory cortex for conditioning, silencing of an ensemble in another side induced no fear response. We also found that mice can find a reward based on the presence or absence of the silencing. Neuronal silencing was stored as working memory. Taken together, we propose that neuronal silencing without explicit activation in the cerebral cortex is enough to elicit a cognitive behavior.fear conditioning | operant conditioning | optogenetics | neuronal inhibition | auditory cortex C ortical neurons exhibit spontaneous activity without explicit external stimuli (1-3), which may not only increase, but also be suppressed, by sensory stimuli (4, 5). For example, auditory stimuli suppress a subset of auditory cortical neurons in a frequency-dependent manner (5). Synaptic inhibition in the cerebral cortex is fundamental for neuronal modulation (6), including gain control (7), response selectivity (8, 9), and synchronized activities (10, 11). Inhibition-based modulations may contribute to stimulusdriven behaviors and associative memories of sensory stimuli (12); however, it remains unclear whether neuronal silencing (i.e., a transient reduction in firing rates from their spontaneous level) by itself can serve as a memory trace and bring about behavioral expressions. In this study, we tested this possibility by optogenetically silencing auditory cortical neurons. ResultsArchaerhodopsin Expression in the Auditory Cortex. To silence excitatory neurons in the auditory cortex, we injected an adenoassociated virus vector expressing archaerhodopsin (Arch) (13) under the control of the calcium/calmodulin-dependent protein kinase II (CaMKII) promoter (AAV-CaMKII-Arch-EYFP) into the auditory cortex of mice (Fig. 1A). Three weeks after virus injection, we observed robust expression of Arch-EYFP in the auditory cortex (Fig. 1B). We obtained whole-cell recordings from Arch-expressing neurons in auditory cortical slices and confirmed that green-light illumination induced an outward current and inhibited action potentials induced by depolarizing current injections ( Fig. 1 C and D). To confirm the inhibitory effect in vivo, we recorded multiunit activities from the auditory cortex of the anesthetized mice. The overall firing rate was reduced by 29.2 ± 7.7% during green-light illumination (eight tetrodes from two mice) (Fig. 1E).Neuronal Silencing Induces Conditio...

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“…First, the multiple trace theory states that in addition to activated cells during the user-defined time windows, non-activated cells would become the engram cells during memory consolidation and retrieval (Nadel et al, 2000 ). The actively inhibited cells involved in other processes of memory (Nomura et al, 2015 ; Vetere et al, 2021 ) should be appreciated during the formation of engram cells. Thus, labeling and manipulating inactivated cells around the activated cells is crucial for studying memory engrams.…”

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“…Because this inhibition is necessary for content-specific memory retrieval success, these inhibited neurons ought to be considered a genuine component of the engram. Similarly, much as re-activating neurons that were active during encoding promotes memory retrieval, re-inhibiting neurons that were actively inhibited during encoding can also promote memory retrieval [ 191 ]. That such neurons are re-inhibited (rather than re-activated) during memory retrieval does not preclude them from being an essential component of the engram (for conceptually related work on inhibitory engrams, see [ 192 , 193 ]).…”