Alarin in different human intestinal epithelial cell types.

Alarin (AL), a new member of the galanin family, has been localized in various CNS regions, mainly in rodents. Among other effects, it modulates food intake. Therefore, we analyzed the immunohistochemical distribution pattern of AL in human intestinal epithelia. Cryosections of 12 human bowel samples were immunohistochemically double-stained for AL and α-defensin 5 (αD; first set). Two further sets of sections were quadruple-stained either (second set) for AL, chromogranin (CG), synaptophysin (SY), and somatostatin (SO) or (third set) for AL, CG, Peptide Y (PY), and 5-hydroxytryptamine (5-HT). Slides were digitized and quantitative analysis of co-localization rates was undertaken. Small bowel: most of AL-positive cells (56%) were αD-positive Paneth cells located within the base of the crypts (first set). In the second set, about 27% of AL-labeled cells were co-reactive for SY and CG, likely representing entero-endocrine cells. In the third set, the largest subpopulation of AL-positive cells was not co-reactive for other markers applied (89%); most of them were likely Paneth cells. Large bowel: co-localization of AL with αD was not detected (first set). In the second set, AL was frequently co-localized with the other three markers applied (68%). In the third set, AL was frequently co-localized with 5-HT and CG (31%) as well as with PY and 5-HT (22%). Due to its presence in various enteroendocrine as well as Paneth cells, AL may be involved in different physiological and pathological processes.

Alarin in cranial autonomic ganglia of human and rat.

Extrinsic and intrinsic sources of the autonomic nervous system contribute to choroidal innervation, thus being responsible for the control of choroidal blood flow, aqueous humor production or intraocular pressure. Neuropeptides are involved in this autonomic control, and amongst those, alarin has been recently introduced. While alarin is present in intrinsic choroidal neurons, it is not clear if these are the only source of neuronal alarin in the choroid. Therefore, we here screened for the presence of alarin in human cranial autonomic ganglia, and also in rat, a species lacking intrinsic choroidal innervation. Cranial autonomic ganglia (i.e., ciliary, CIL; pterygopalatine, PPG; superior cervical, SCG; trigeminal ganglion, TRI) of human and rat were prepared for immunohistochemistry against murine and human alarin, respectively. Additionally, double staining experiments for alarin and choline acetyltransferase (ChAT), tyrosine hydroxilase (TH), substance P (SP) were performed in human and rat ganglia for unequivocal identification of ganglia. For documentation, confocal laser scanning microscopy was used, while quantitative RT-PCR was applied to confirm immunohistochemical data and to detect alarin mRNA expression. In humans, alarin-like immunoreactivity (alarin-LI) was detected in intrinsic neurons and nerve fibers of the choroidal stroma, but was lacking in CIL, PPG, SCG and TRI. In rat, alarin-LI was detected in only a minority of cranial autonomic ganglia (CIL: 3.5%; PPG: 0.4%; SCG: 1.9%; TRI: 1%). qRT-PCR confirmed the low expression level of alarin mRNA in rat ganglia. Since alarin-LI was absent in human cranial autonomic ganglia, and only present in few neurons of rat cranial autonomic ganglia, we consider it of low impact in extrinsic ocular innervation in those species. Nevertheless, it seems important for intrinsic choroidal innervation in humans, where it could serve as intrinsic choroidal marker.

Galanin- but not galanin-like peptide-containing axon terminals innervate hypophysiotropic TRH-synthesizing neurons in the hypothalamic paraventricular nucleus.

Galanin and galanin-like peptide (GALP) are both orexigenic peptides involved in the regulation of food intake and energy metabolism. To determine whether these peptides may directly influence the hypophysiotropic thyrotropin-releasing hormone (TRH)-synthesizing neurons, double-labeling immunocytochemistry was performed at light and electron microscopic levels using antisera against proTRH, galanin and GALP. Galanin-IR axons densely innervated all of the major parvocellular subdivisions of the PVN where proTRH neurons were identified. The periventricular and anterior parvocellular subdivisions exhibited a prominent network of galaninergic nerve fibers, while the density of fibers was less intense in the medial parvocellular subdivision. Galanin-immunoreactive (IR) axon varicosities were juxtaposed to the majority of TRH-synthesizing neurons in the anterior, medial and periventricular subdivisions of the PVN. Ultrastucturally, galanin-IR nerve terminals established symmetric type synapses with the perikarya of proTRH-IR neurons, suggesting an inhibitory nature of these contacts. In contrast, GALP immunoreactive fibers and nerve terminals concentrated primarily in the anterior parvocellular subdivision of the PVN and were found in association with only few proTRH-IR neurons in the periventricular and medial parvocellular subdivisions. In conclusion, the dense innervation of TRH neurons in all subdivisions of the PVN by galanin-IR axons indicates that galanin may be involved in the central regulation of the hypothalamic-pituitary-thyroid axis. In contrast, the relative paucity of GALP-containing axons in juxtapsoition to TRH neurons in the medial and periventricular parvocellular subdivisions of the PVN, the origin of hypophysiotropic TRH neurons, makes it unlikely that GALP similarly exerts direct regulatory effects on hypophysiotropic TRH neurons.