Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to investigate the effects of locally manipulated neurotransmitter and signaling pathways in awake, behaving animals. Moreover, the stereotaxic implantation of electrodes for electrophysiological stimulation and recording studies has been instrumental to our current understanding of neuroplasticity and brain networks in behaving animals. Ever-increasing knowledge about optimizing surgical techniques in rodents 1-4 , public awareness concerning animal welfare issues and stringent legislation (e.g., the 2010 European Union Directive on the use of laboratory animals 5 ) prompted us to refine these surgical procedures, particularly with respect to implementing new procedures for oxygen supplementation and the continuous monitoring of blood oxygenation and heart rate levels during the surgery as well as introducing a standardized protocol for post-surgical care. Our observations indicate that these modifications resulted in an increased survival rate and an improvement in the general condition of the animals after surgery (e.g. less weight loss and a more active animal). This video presentation will show the general procedures involved in this type of stereotaxic surgery with special attention to our several modifications. We will illustrate these surgical procedures in rats, but it is also possible to perform this type of surgery in mice or other small laboratory animals by using special adaptors for the stereotaxic apparatus 6 . Video LinkThe video component of this article can be found at http://www.jove.com/video/3528/ Protocol Note: Antiseptic techniques should be employed during the whole procedure. All the instruments and materials (cotton-tipped swabs, gauze, etc.) that will be used during the surgery should be sterilized by autoclaving. A surgical mask, hair bonnet and sterile gloves should be worn. The working area and the stereotaxic apparatus should be cleaned thoroughly, and disinfected with a 70% ethanol solution. 2. Place the cannula in its support and check if it is straight. 3. Turn on the gas system -mixture of ambient air and oxygen (30-35% of total flow should be oxygen). 4. Weigh the rat and administer the anesthetic. We are using a mixture of ketamine (37.5 mg/kg) and dexmedetomidine (0.25 mg/kg) injected subcutaneously. For different anesthesia protocols, see Flecknell 4 and Hellebrekers et al. 7 . 5. After the rat lost consciousness, shave the head area going from the ears to just in between the eyes with an electric razor. 6. Place the rat on the heating pad, with its nose in front of the air tubing. Use an oximeter to ensure that the rat has an adequate blood oxygenation level (should not drop <90%). Please follow the manufacturer's instructions for proper use of equipment. 7. Apply eye cream (Duratears Z, Alcon) on both corneas to avoid dehydration. 8. Check the rat's reflexes (tail reflex or toe-pinch reflex, as demonstrated in Walantu...

It is well established that arousal-induced memory enhancement requires noradrenergic activation of the basolateral complex of the amygdala (BLA) and modulatory influences on information storage processes in its many target regions. While this concept is well accepted, the molecular basis of such BLA effects on neural plasticity changes within other brain regions remains to be elucidated. The present study investigated whether noradrenergic activation of the BLA after object recognition training induces chromatin remodeling through histone post-translational modifications in the insular cortex (IC), a brain region that is importantly involved in object recognition memory. Male Sprague—Dawley rats were trained on an object recognition task, followed immediately by bilateral microinfusions of norepinephrine (1.0 μg) or saline administered into the BLA. Saline-treated control rats exhibited poor 24-h retention, whereas norepinephrine treatment induced robust 24-h object recognition memory. Most importantly, this memory-enhancing dose of norepinephrine induced a global reduction in the acetylation levels of histone H3 at lysine 14, H2B and H4 in the IC 1 h later, whereas it had no effect on the phosphorylation of histone H3 at serine 10 or tri-methylation of histone H3 at lysine 27. Norepinephrine administered into the BLA of non-trained control rats did not induce any changes in the histone marks investigated in this study. These findings indicate that noradrenergic activation of the BLA induces training-specific effects on chromatin remodeling mechanisms, and presumably gene transcription, in its target regions, which may contribute to the understanding of the molecular mechanisms of stress and emotional arousal effects on memory consolidation.

Histone posttranslational modifications (PTMs), by their action on the chromatin state, play a central role in the regulation of gene expression. The discovery that some PTMs in the brain are dynamically regulated by experience and environmental factors makes them an important subject for the study of plasticity changes in learning and memory, addiction, and psychiatric disorders. Current histone isolation protocols, however, require large amounts of tissue, which limits their application for analyzing small tissue samples from a specific brain region. We describe here a step-by-step protocol for histone extraction and isolation from 1 mm(3) of tissue from brain punches, which allows reproducible and reliable results for histone PTM identification and quantification without losing anatomical precision. © 2016 by John Wiley & Sons, Inc.