No 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.

The prelimbic-medial orbital areas (PL/MO) of the prefrontal cortex are connected to the medial part of the subthalamic nucleus (STN) through a direct projection and an indirect circuit that involves the core of the nucleus accumbens (NAcc) and the ventral pallidum (VP). In the present study, the influence of the PL/MO on the discharge of STN cells has been characterized. The major pattern of the responses observed after stimulation of PL/MO consisted of two excitatory peaks often separated by a brief inhibitory period. The early excitation was most likely to be caused by the activation of direct cortical inputs because its latency matches the conduction time of the prefrontal STN projections. The late excitation resulted from the activation of the indirect PL/MO-STN pathway that operates through a disinhibitory process. Indeed, the late excitation was no longer observed after acute blockade of the glutamatergic corticostriatal transmission by CNQX application into the NAcc. A similar effect was obtained after the blockade of the GABAergic striatopallidal transmission by bicuculline application into the VP. Finally, the brief inhibition that followed the early excitation was likely to result from the activation of a feedback inhibitory loop through VP because this inhibition was no longer observed after the blockade of STN inputs by CNQX application into the VP. This study further indicates the implication of STN in prefrontal basal ganglia circuits and underlines that in addition to a direct excitatory input, medial STN receives an indirect excitatory influence from PL/MO through an NAcc-VP-STN disinhibitory circuit. Key words: basal ganglia circuits; prefrontal cortex; subthalamic nucleus; ventral striatum; nucleus accumbens; ventral pallidum; in vivo single unit recordings; ratThe subthalamic nucleus (STN) is a major component of the basal ganglia, and its critical role in the control of movement is well established. Pathological damage to the STN in humans or STN lesions in monkeys induce hemiballism (Whittier, 1947;Carpenter et al., 1950;Crossman, 1987). In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated monkeys, an animal model of Parkinson's disease, abnormal activity of STN neurons has been observed, and lesions or high-frequency stimulation of the STN ameliorates akinesia and rigidity (Bergman et al., 1990;Benazzouz et al., 1993). High-frequency stimulation of the STN is now successf ully applied to improve akinesia and rigidity in Parkinsonian patients (Limousin et al., 1995).In current models of the basal ganglia circuitry, the striatum and the STN are the two major structures through which cortical signals are transmitted to the output structures of the basal ganglia, i.e., the substantia nigra pars reticulata (SN R) and the internal segment of the globus pallidus (GPi) (Alexander and Crutcher, 1990; Parent and Hazrati, 1995a,b). Indeed, both the striatum and the STN receive direct excitatory cortical inputs and send projections to the SN R and the GPi. Because the projection neurons of the stri...