Background

Nucleus accumbens (NAc) is part of the basal ganglia located in the rostroventral part of the striatum. Specifically, NAc is part of a structure that we call ventral striatum, which also comprises the olfactory tubercle. It is well known the role of NAc in the dopaminergic mesolimbic pathway, which regulates motivational behavior and is responsible for emotional and contextual behaviors (Baik, 2013). NAc has already been implicated in psychiatric disorders (e.g., depression), drug addiction, obesity, and chronic pain, which show the relevance in studying the neurobiology of this brain structure (Nestler and Carlezon, 2006; Volkow et al., 2011; Volkow and Morales, 2015; Brandão et al., 2019; Serafini et al., 2020).

Based on this, it is important to have a precise technique describing how to microdissect the NAc, sampling this structure for molecular analysis. The punch technique (Palkovits, 1983), for example, allow us to microdissect brain sections (including NAc) with a hollow needle and is widely used in neurobiological laboratories. Advantages of punch technique are low cost and fast sampling. Disadvantage of using this technique is the limited accuracy, once the experimenter is restricted to the cylindrical shape of the punch tool and because the experimenter must punch fresh tissue, which is hard to identify some surrounding neuroanatomic structures of the NAc, such as the ventral pallidum.

Because of this, depending on the molecular analysis and the objective, it is preferable using a more accurate technique, such as laser-assisted microdissection (or laser capture microdissection). Laser-assisted microdissection technique is a method to microdissect tissue under direct microscopic visualization (Espina et al., 2006). Advantages of using this technique are the high accuracy on delineation of the NAc and the capacity of sampling smaller areas (e.g., NAc core or shell) or even cell populations. Disadvantages of this technique are the long time until sampling the tissue and the high cost to perform, requiring an entire facility with microscope and laser equipment.

Interestingly, Franck et al. (2013) described the parafilm-assisted microdissection (PAM) as a sampling method, a technique which involves the microdissection of tissue sections mounted on parafilm-covered glass slides. This group described the application of this technique for prostate cancer (Quanico et al., 2015), ovarian tissue (Delcourt et al., 2017) and for specific brain areas such as cerebellum, hypothalamus, and hippocampus (Franck et al., 2013; Quanico et al., 2017a and 2017b; Delcourt et al., 2018). The PAM technique can be easily performed, once it uses low-cost materials and allows high accuracy on delineating the target brain area. Disadvantage of this technique is the limitation in microdissection smaller structures, such as subthalamic nucleus, for example.

In this manuscript, we will describe the whole process, from the brain storage after euthanasia to molecular analysis, of using the PAM technique to specifically microdissect the rodent NAc, also describing how to stain the brain slices to easily identify the anatomical limits of the NAc. It is worth mentioning that, as above-mentioned, this protocol can be used to microdissect a great range of brain regions, depending on the size and shape. Tissue obtained from this technique can be used for molecular analyzes such as real-time PCR, Western Blot, and RNA-seq.