fNIRS technology applied on animals: a study on sheep

The lack of data concerning the organization of the central nervous system of the animals farmed for milk and meat produc- tion is a potential handicap for the study of cognition processes, important for the assessment of animal welfare, and a gap in comparative neuroscience. We conducted several trials in order to study the functional organization of the cerebral cortex of free-moving sheep through the non-invasive application of functional near-infrared spectroscopy (fNIRS), a developing tech- nology that has started to be applied in animals (Muehlemann et al., 2011).We intended to record the oxygen consumption of selected areas of the brain involved in the performance of behavioural motor and associative tasks and to assess the brain activity when animals were anticipating either a positive or negative event. We used the continuous-wave fNIRS OxyPrem device and recorded the data with Tubis software (version 4.5). Here we present the fNIRS results of the pilot phase that was conducted on three sheep undergoing to a series of stimuli for 20 s, each followed by a 20 s interval. The stimuli applied were: 1) auditory, consisting in a neutral sound repeated 8 times; 2) visual, involving the application of the Finnoff light intermittently on the pupil, repeated 4 times per eye and 3) physical, entailing the application of a pressure of 250 mmHg with a tourniquet on the forelimb, 4 times per limb. We repeated the pro- tocol recording the brain activity with electroencephalography (EEG). In addition, we set up an experiment to assess the brain activity when animals were anticipating either a positive or negative event. We trained eight sheep to anticipate two events with a supposed different value via classical conditioning. Two pairs of audio-visual stimuli anticipated either the presence of feed (positive event) or the absence of it (aversive event) in a testing area hidden behind screens. On test day, each sheep performed two sessions of six trials. A session consisted in 3 positive and 3 negative trials randomly distributed (always starting with a positive). During the task sheep were wearing the fNIRS device on the head. We focused on the brain activity during the 20 s of latency between submission to stimuli and events. Furthermore, we analysed the behaviour of the sheep during the anticipation period using validated ethograms (Reefmann et al., 2009; Boissy et al., 2011). fNIRS represents an innovative non-invasive method to conduct more objective assessments of animal behaviour and helps to improve the evaluation of animal welfare. The data will allow the identification of the cerebral areas involved in the physio- logical regulation of complex and integrated behaviours with different emotional valences. Combining the exploration of the neural substrates underlying cognitive functions with existing behavioural and physiological measures will strengthen knowledge of how animals perceive different environmental situations. Validation of the data will also promote the use of large herbivores\u2019 brains as suitable scientific models in neuroscience