Exploring the sense of number in zebrafish: a neurobiological approach

The ability to deal with continuous (spatial or temporal) and discrete quantity (numerosity) developed from an evolutionarily conserved system for approximating non-symbolic numerical magnitude, which has been documented in a variety of species, including fish. In fish numerosity discrimination has been documented using spontaneous choice tests and operant conditioning procedures, however little is knowns about the neural correlates of this ability. By combining a habituation/dishabituation behavioral paradigm with molecular biology assays, we have recently identified part of the neural network associated with quantity discrimination in adult zebrafish brain. Zebrafish were habituated to groups of 3 or 9 small red dots for four consecutive days. During this phase, the dots changed in density, position and size (thus preventing the habituation of fish to a specific configuration of the stimulus though maintaining their numerousness and total surface area. During dishabituation, zebrafish faced a change (i) in number (from 3 to 9 dots or vice versa, with the same overall surface), or (ii) in shape (3 or 9 red squares instead of 3 or 9 dots with the same overall surface), or (iii) in size (with the same shape and number). A control group was tested with the same stimuli as during the habituation. Thirty minutes after the dishabituation test, zebrafish were sacrificed, their brains were collected and dissected to quantify the change in the expression levels of c-fos and egr-1 by quantitative polymerase chain reaction (qPCR) or probed with egr-1 in situ hybridization assays to identify the positional identity of neuronal correlates discriminating changes in quantity (number, size) or shape. Results showed an involvement of the retina and optic tectum in the encoding of continuous magnitude (e.g., a change in stimulus size). We also found a role of the habenula and the preglomerular complex, and of the caudal regions of the dorso-lateral and dorso-central pallium, in the encoding of discrete magnitude (e.g. change in numerosity). A response to shape discrimination was observed in the most rostral part of the dorso-central pallium. Results suggest an early involvement of thalamic and tectal areas for encoding of continuous quantity, and of more pallial (via talamac nuclei) regions for discrete quantity.