Abstract
Precise identification of neuronal populations is a major challenge in neuroscience. In the striatum, more than 95% of neurons are GABAergic medium-sized spiny neurons (MSNs), which form two intermingled populations distinguished by their projections and protein content. Those expressing dopamine D1-receptors (D1Rs) project preferentially to the substantia nigra pars reticulata (SNr), whereas those expressing dopamine D2- receptors (D2Rs) project preferentially to the lateral part of the globus pallidus (LGP). The degree of segregation of these populations has been a continuous subject of debate, and the recent introduction of bacterial artificial chromosome (BAC) transgenic mice expressing fluorescent proteins driven by specific promoters was a major progress to facilitate striatal neuron identification. However, the fraction of MSNs labeled in these mice has been recently called into question, casting doubt on the generality of results obtained with such approaches. Here, we performed an in-depth quantitative analysis of striatal neurons in drd1a-EGFP and drd2-EGFP mice. We first quantified neuronal and non-neuronal populations in the striatum, based on nuclear staining with TO-PRO-3, and immunolabeling for NeuN, DARPP-32 (dopamine- and cAMP-regulated phosphoprotein Mr∼32,000), and various markers for interneurons. TO-PRO-3 staining was sufficient to identify MSNs by their typical nuclear morphology and, with a good probability, interneuron populations. In drd1a-EGFP/drd2-EGFP double transgenic mice all MSNs expressed EGFP, which was driven in about half of them by drd1a promoter. Retrograde labeling showed that all MSNs projecting to the SNr expressed D1R and very few D2R (<1%). In contrast, our results were compatible with the existence of some D1R-EGFP-expressing fibers giving off terminals in the LGP. Thus, our study shows that nuclear staining is a simple method for identifying MSNs and other striatal neurons. It also unambiguously confirms the degree of segregation of MSNs in the mouse striatum and allows the full exploitation of results obtained with BAC-transgenic mice.
Highlights
Basal ganglia form a complex neural network involved in the selection and execution of action through interactions with multiple brain areas that process sensorimotor, emotional and cognitive information [1]
We noticed that nuclear staining with TO-PRO-3 was heterogeneous in mouse striatal sections, as in other brain regions, and we investigated whether this heterogeneity matched with known cellular types
We first characterized the staining pattern of TOPRO-3 in cells labeled with DARPP-32, a regulatory protein found in cytoplasm and nuclei of Medium-sized spiny neurons (MSNs) [25,26]
Summary
Basal ganglia form a complex neural network involved in the selection and execution of action through interactions with multiple brain areas that process sensorimotor, emotional and cognitive information [1]. MSNs receive excitatory glutamatergic inputs from the cerebral cortex and the thalamus, and a modulatory dopaminergic innervation from the midbrain. They belong to two intermingled subpopulations distinguished by their projections and protein expression patterns. MSNs expressing dopamine D1-receptors (D1Rs), dynorphin and substance P, project to the substantia nigra pars reticulata (SNr) and entopeduncular nucleus (direct striatonigral pathway), while MSNs expressing dopamine D2-receptors (D2Rs) and enkephalin, project to the lateral part of the globus pallidus (LGP) (indirect striatopallidal pathway) [5,6,7] These two subpopulations are homogenously distributed throughout the striatum, and are known to have opposite behavioral effects, as they are coupled to output pathways with opposing properties [8]. The exact degree of segregation between these two types of MSNs has been much disputed
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call