Neuropeptide co-expression in hypothalamic kisspeptin neurons of laboratory animals and the human

Abstract

Hypothalamic peptidergic neurons using kisspeptin (KP) and its co-transmitters for communication are critically involved in the regulation of mammalian reproduction and puberty. This article provides an overview of neuropeptides present in KP neurons, with a focus on the human species. Immunohistochemical studies reveal that large subsets of human KP neurons synthesize neurokinin B, as also shown in laboratory animals. In contrast, dynorphin described in KP neurons of rodents and sheep is found rarely in KP cells of human males and postmenopausal females. Similarly, galanin is detectable in mouse, but not human, KP cells, whereas substance P, cocaine- and amphetamine-regulated transcript and proenkephalin-derived opioids are expressed in varying subsets of KP neurons in humans, but not reported in ARC of other species. Human KP neurons do not contain neurotensin, cholecystokinin, proopiomelanocortin-derivatives, agouti-related protein, neuropeptide Y, somatostatin or tyrosine hydroxylase (dopamine). These data identify the possible co-transmitters of human KP cells. Neurochemical properties distinct from those of laboratory species indicate that humans use considerably different neurotransmitter mechanisms to regulate fertility.

Keywords: CART; dynorphin; hypothalamus; kisspeptin; neurokinin B; reproduction; substance P.

Figure 1

Results of immunofluorescent studies to characterize the neuropeptide phenotype of human KP neurons in the Inf. (A,B) The highest numbers of KP and NKB neurons and colocalization percentages can be observed in the Inf of postmenopausal women (A). Compare to the weaker labeling of the Inf from a 67 year-old men in (B). Note that the Inf contains many single-labeled axons (red and green), in addition to dual-labeled ones (yellow) in both sexes. (C) Unlike KP neurons of laboratory animals, the majority of human KP neurons do not contain Dyn A immunoreactivity. A rare case of dual-labeled KP/Dyn A axon (arrows) is shown at higher power in the inset. (D) KP neurons often contain SP immunoreactivity in postmenopausal women. White cells labeled by arrowheads correspond to KP/NKB/SP triple-phenotype neurons (Hrabovszky et al., 2013). (E,F) Similarly, CART is co-expressed in large subsets of KP and NKB neurons in postmenopausal women (Skrapits et al., 2014). Arrowhead in (E) and the two arrows in (F) point to a triple-labeled perikaryon and two axon varicosities, respectively. (G) The majority of neuropeptides tested in this study (see Figure 2), including galanin (blue), showed no colocalization with KP or NKB. (H–J) One exception was pENK which occurred in subsets of NKB-IR and KP-IR perikarya and fibers. Arrowhead in (H) points to a perikaryon, whereas arrows in (I,J) label axon varicosities that exhibit triple-neuropeptide phenotype (KP/NKB/pENK). The results shown in (H,I) vs. (J) were obtained with two different pENK antibodies (see text) to serve as positive control. (Note that the original color of the fluorochromes was changed so that KP is illustrated consistently in red). Scale bars: 40 μm in (A,B,D,E,G,H) and 12 μm in (C,F,I,J).

Figure 2

Summary of neuropeptide/neurotransmitter co-expression data in KP neurons of laboratory animals and the human. AGRP, agouti-related protein; αMSH, α-melanocyte stimulating hormone; ARC, arcuate nucleus; AVPV, anterior periventricular nucleus; CART, cocaine- and amphetamine regulated transcript; CCK, cholecystokinin; Dyn, dynorphin; GAD-67, glutamic acid decarboxylase-67; GAL, galanin; Inf, infundibular nucleus; ISH, in situ hybridization; KP, kisspeptin; mENK, met-enkephalin; NKB, neurokinin B; NPY, neuropeptide Y; NT, neurotensin; pENK, proenkephalin; POMC, proopiomelanocortin; RP3V, rostral periventricular area of the third ventricle; SP, substance P; SS, somatostatin; TH, tyrosine hydroxylase; vGluT2, type-2 vesicular glutamate transporter.