Ultrasound-guided cerebrospinal fluid collection from rats

•We describe an ultrasound-guided method for CSF collection from cisterna magna in rat.•This method allows accurate puncture with help from the real-time ultrasound images, yields a considerable amount of blood-free CSF (100–200 μL), and causes minimal injury.•Using this method, we demonstrate that repeated morphine administration significantly decreases CSF β-endorphin levels in rats.•This method can facilitate the study of neurobiology, pharmacology and behaviour.