Subcellular distribution of histamine, GABA and galanin in tuberomamillary neurons in vitro

Abstract

Histamine acts as a neurotransmitter in the brain and regulates e.g. sleep, hibernation, vigilance, and release of several other transmitters. All histaminergic neurons are found in the tuberomamillary nucleus (TM), and send axons to almost all parts of the CNS. Despite the obvious importance of these neurons, their development, transmitter storage, and compartmentalization of cotransmitters are poorly known. Histaminergic neurons from fetal rat hypothalamus were studied in primary explant cultures and analyzed by confocal microscopy. Most histaminergic neurons were oval in shape, but round and triangular ones were also found. The average size of the 212 analyzed neurons was 19.2 μm (length), 12.5 μm (width) and 11.7 μm (thickness). The cells possessed two to five microtubule-associated protein (MAP2) positive processes, putative dendrites, and in general one MAP2-negative thin process, a putative axon. Granular histamine-immunoreactivity was found in the cell bodies, axons, and dendrites. In tuberomamillary neurons, most histamine-containing structures displayed immunoreactivity for vesicular monoamine transporter 2 (VMAT2), indicating that the two markers may coexist in the same structures. Lack of VMAT2 in some histamine-immunoreactive structures indicates that another transporter for histamine may exist. In the same neurons, γ-aminobutyric acid (GABA)-immunoreactivity was found in structures, distinct from those containing histamine, indicating that the two transmitters may be differentially localized, regulated and released. Galanin-immunoreactivity in the cultured tuberomamillary neurons was partially located in the same structures as VMAT2. The results suggest that histamine and GABA, the two principal transmitters of tuberomamillary neurons, are not costored in the same structures in tuberomamillary neurons.