Immunohistochemical localization of vesicular glutamate transporter 2 (vGluT2) in the central nervous system of the pigeon (Columba livia)

Abstract

Glutamatergic neurons are distributed widely in the telencephalic pallium of birds, but their targets are unknown. In the present study, a polyclonal antibody was produced against pigeon vesicular glutamate transporter 2 (vGluT2) and was used for Western blot and immunohistochemistry to detect projection targets of glutamatergic neurons in the pigeon central nervous system. The molecular weight of vGluT2 was measured at 65 kDa. vGluT2 immunoreactivity was observed in neuropil, but not in neuronal cell bodies or glia. Immunoreactive neuropil generally appeared homogeneous, but fine granules or puncta were found in many areas or nuclei. The telencephalon showed strong immunoreactivity, except for the globus pallidus. In particular, the mesopallium and hippocampal formation revealed the most intense immunostaining. In the diencephalon, vGluT2 immunoreactivity was intense in the dorsal thalamus and hypothalamus. In the midbrain, strong immunostaining was seen in the periaqueductal gray, the dorsal part of the lateral mesencephalic nucleus, and the isthmo-optic nucleus. The optic tectum showed moderate immunoreactivity. In the cerebellar cortex, glomeruli in the granular layer were intensely immunoreactive, and the molecular layer showed intensely homogeneous immunostaining. In the caudal brainstem, the cochlear magnocellular and angular nuclei showed strongly immunoreactive puncta around neuronal cell bodies. In the spinal cord, the dorsal horn revealed moderate immunoreactivity and the marginal nucleus was strongly immunoreactive. Ultrastructural observations revealed that vGluT2 immunoreactivity is localized in asymmetric, presynaptic terminals. The present results indicate an extensive distribution of glutamatergic projections and circuits in the avian central nervous system.

Proposed homology of the dorsomedial subdivision and V‐shaped layer of the avian hippocampus to Ammon's horn and dentate gyrus, respectively

“…In situ hybridization studies using a vGluT2 probe morphologically demonstrated the presence of a large number of glutamatergic neurons in HF of pigeons and zebra finches, especially apparent in neurons of the V‐shaped layer (Islam and Atoji, ; Karim et al, ; present study). Immunohistochemically, vGluT2 protein has been ultrastrucutrally demonstrated to be localized in presynaptic boutons of asymmetric synapses in the pigeon brain (Atoji, ). Considering the localization of zinc‐containing presynaptic boutons described above, vGluT2 probably colocalizes in zinc‐containing axon terminals.…”

Expression of ionotropic glutamate receptors, AMPA, kainite and NMDA, in the pigeon retina

“…The localization of GluA1 neurons in the optic tectum largely overlaps with that of vGluT2 immunoreactive axon terminals (Atoji, 2011). In contrast, the dorsal thalamic region weakly expresses GluA2 mRNA but not GluA1 mRNA (Ottiger et al, 1995;Islam and Atoji, 2008), but the thalamic region revealed intense vGluT2 immunoreactivity (Atoji, 2011). The difference of mRNA expression of GluA1 and GluA2 between the optic tectum and dorsal thalamic nuclei is unknown.…”

“…GluA1 is immunohistochemically localized in neurons of layers 2e7, 9, and 10 in the pigeon optic tectum (Theiss et al, 1998). The localization of GluA1 neurons in the optic tectum largely overlaps with that of vGluT2 immunoreactive axon terminals (Atoji, 2011). In contrast, the dorsal thalamic region weakly expresses GluA2 mRNA but not GluA1 mRNA (Ottiger et al, 1995;Islam and Atoji, 2008), but the thalamic region revealed intense vGluT2 immunoreactivity (Atoji, 2011).…”

Microconnectomics of the pretectum and ventral thalamus in the chicken (Gallus gallus)

“…The GLv, LM, and GT receive topographic retinal projections from different retinal ganglion cell populations (Crossland and Uchwat, ; Gamlin and Cohen, ; Wylie et al, ), which may release glutamate in all these structures (Guo et al, ; Atoji, ). At the same time, AMPA receptors GluR1, GluR2/3, and GluR4 are expressed in the GT, LM, and GLv (Pires and Britto, ).…”