Oxy-haemoglobin (HbO2) and deoxy-haemoglobin (Hb) are chromophores present in biological tissues. Near infrared spectroscopy (NIRS) is a non-invasive methodology based on the low extinction coefficient of tissue in the near infrared region. NIRS can be used to measure changes in the concentration of these chromophores, i.e., haemoglobin, in muscular tissue. In the present work, NIRS has been used for the non-invasive monitoring of HbO2, Hb, and blood volume (V: representing total haemoglobin, i.e., HbO2+Hb) in vivo in the whole rat brain. This has been performed by means of prototype instrumentation based on optic fibre probes placed in contact with the head of anaesthetised rats held in a stereotaxic frame. A preliminary test of the instrument has been performed on human muscle, i.e., lateral gastrocnemius, in order to evaluate the ability of the instrument to detect oxygenation changes. Afterwards, the effects of pharmacological treatments, such as systemic amphetamine and nicotine treatments on the CNS have been detected.