Scale bars = 1 mm in (A); 200 m in (BCH). Open in a separate window Figure 5 Distribution of axonal fibers form neocortical GABAergic projection neurons in case mRT210. markers revealed the majority of corticostriatal GABAergic projection neurons were positive for somatostatin (SS)-immunoreactivity. In contrast, corticothalamic GABAergic projection neurons were not recognized by representative neurochemical markers for GABAergic neurons. These findings suggest that corticofugal GABAergic projection neurons are heterogeneous in terms of their neurochemical properties and target nuclei, and provide axonal innervations mainly to the nuclei in the basal ganglia. = 18). We confirmed the injection sites were relatively larger (0.6C1.0 mm) and were confined only in frontal cortex (Figures ?(Figures33C5) in 10 mice. These 10 mice were chosen for further analysis. Neurons labeled by AAV-Ef1a-DIO-hChR2-mCherry were found within the injection sites such as the dorsolateral, lateral, and ventral orbital, and main and secondary motor cortices. In addition, mCherry-immunoreactive fibers were found throughout the rostral-caudal extent of the brain. The figures and intensities of mCherry-immunoreactive fibers were diverse across animals, presumably because of the different sizes of injection sites. In contrast, the patterns of mCherry-immunoreactive fibers were comparable across all animals injected, with some differences (Table ?(Table2).2). Abundant mCherry-immunoreactive fibers with en passant synapses were observed in the cerebral cortex, basal ganglia and thalamus (Physique ?(Figure6).6). Additionally mCherry-immunoreactive fibers appeared in the amygdala (Physique ?(Figure6C)6C) in three cases, and in the dorsal raphe nucleus in one case (Figure ?(Physique5C).5C). The details of axonal distribution are briefly summarized below. Open in a separate window Physique 3 Distribution of axonal fibers form neocortical GABAergic projection neurons in case mRT217. Low (A) and high (BCH) magnification lightfield photomicrographs showing mCherry-immunoreactive fibers in selected sections after the injection of AAV-Ef1a-DIO-hChR2-mCherry mainly in the orbital cortex (A1). The mCherry-immunoreactive fibers were found in the Tg dorsal peduncular and infralimbic cortex (A2,B), in the lateral accumbens shell and olfactory tubercle (A3,C), in the ventral caudate putamen (A4CA7,DCG), in the ventral pallidam (A6,F), in the lateral globus pallidus (A7,G), and in the mediodorsal thalamic nucleus (A8,H). Note that mCherry-immunoreactive fibers were also found in the contralateral mediodorsal thalamic nucleus. The dashed collection indicates the midline of the brain. aca, anterior commissure, anterior part; acp, anterior commissure, posterior part; CPu, caudate putamen; Cx, neocortex; fmi, forceps minor of the corpus callosum; ic, internal capsule; LAchSh, lateral accumbens shell; LGP, lateral globus pallidus; LV, lateral ventricle; Tu, olfactory tubercle; VP, ventral pallidum. Scale bars = 1 mm in (A); 200 m in (BCH). Open in a separate window Figure 5 Distribution of axonal fibers form neocortical GABAergic projection neurons in case mRT210. Low (A) and high (B,C) magnification lightfield photomicrographs showing mCherry-immunoreactive fibers in selected sections after the injection of AAV-Ef1a-DIO-hChR2-mCherry mainly in the orbital cortex (A1). The mCherry-immunoreactive fibers were found in the LGP Pozanicline (A2,B) and in the dorsal raphe nucleus (A3,C). Scale bars = 1 mm in (A); 200 m in (B); 30 m in (C). Table 2 Relative density of GABAergic fibers in various brain structures from representative injections. = 8). We confirmed the injection sites were mainly confined in the ventral part of CPu and LGP. Injections of FB into the ventral part of CPu resulted in numerous FB-labeled neurons mainly in layers 5 and 2/3 of the frontal cortex, consistent with the previous report in Gabbott et al. (2005), and very occasionally FB- and GFP-double labeled neurons in the frontal cortex (Figure ?(Figure7).7). Interestingly, FB- and GFP-double-labeled neurons did not mingle with numerous FB-labeled neurons. Rather FB- Pozanicline and GFP-double-labeled neurons were found in the regions with sparsely FB-labeled neurons. The FB- and GFP-double labeled neurons were observed mainly in infragranular layers, but also in layer 2/3 and the subcortical white matter. Next, we further examined the neurochemical properties of FB- and GFP-double labeled neurons using antibodies for CR, nNOS, NPY, PV, SS, and VIP. Triple labeling demonstrated that Pozanicline most of FB- and GFP-double-labeled neurons exhibited SS-immunoreactivity (53%: 41/78).