Plasticity of Lh cells caused by cell proliferation and recruitment of existing cells

Abstract

Luteinizing hormone (Lh) and follicle-stimulating hormone (Fsh) control reproduction in vertebrates. Using a transgenic line of medaka, in which green fluorescent protein expression is controlled by the endogenouslhbpromotor, we studied development and plasticity of Lh cells, comparing juveniles and adults of both genders. Confocal imaging and 3D reconstruction revealed hypertrophy and hyperplasia of Lh cells in both genders from juvenile to adult stages. We show that Lh cell hyperplasia may be caused by recruitment of existing pituitary cells that start to producelhb, as evidenced by time lapse recordings of primary pituitary cell cultures, and/or through Lh cell proliferation, demonstrated through a combination of 5-bromo-2′-deoxyuridine incubation experiments and proliferating cell nuclear antigen staining. Proliferating Lh cells do not belong to the classical type of multipotent stem cells, as they do not stain with anti-sox2. Estradiol exposurein vivoincreased pituitary cell proliferation, particularly Lh cells, whereas pituitarylhbandgpaexpression levels decreased. RNA-seq andin situhybridization showed that Lh cells express two estrogen receptors,esr1andesr2b, and the aromatase genecyp19a1b, suggesting a direct effect of estradiol, and possibly androgens, on Lh cell proliferation. In conclusion, our study reveals a high degree of plasticity in the medaka Lh cell population, resulting from a combination of recruitment and cell proliferation.

Plasticity in medaka gonadotropes via cell proliferation and phenotypic conversion

“…Contrary to what has been previously described for Lh cells (Hildahl et al 2012), we did not observe any Fsh cell outside of the pituitary in medaka. We then observed that similar to in Lh cells (Fontaine et al 2019), the number of Fsh cells as well as the percentage of cells they represent in the pituitary increase between juvenile and adult stages. In addition, we demonstrate that the cell volume is also increasing between juvenile and adult stages, which is in agreement with the previous observation where Lh cell size was also observed to increase.…”

“…Counting of Fsh cells was performed blindly using Cell Profiler software (v2.1.0 (Carpenter et al 2006)) as described in Fontaine et al (2019), from eight to nine animals from each sex and stage. Double-labeled cells (BrdU/hrGfpII or BrdU/DsRed2) after steroid and BrdU treatments were manually counted using Fiji software and cell-counter plugin.…”

“…Previously, several studies conducted on Lh cells in medaka have explored and investigated basic parameters including morphology, ontogeny and regulation of Lh cells. In medaka, Lh cells have been found to participate in the plasticity of the pituitary during puberty through hypertrophy and estrogen-sensitive hyperplasia during puberty (Fontaine et al 2019). Lh cells have also been shown to make neuron-like projections allowing homotypic networks (Grønlien et al 2019), to express gnrh receptors (gnrhr), and to respond to gnrh stimuli by increasing their action potential frequency and intracellular calcium concentration (Strandabo et al 2013.…”

Gnrh1-induced responses are indirect in female medaka Fsh cells

“…Fsh cells were located in the median part of the proximalis pars distalis (PPD). Homozygous tg( fshb :DsRed2) fish were crossed with the previously established tg( lhb :hrGfpII) line (Figure 1E-H) established by Hildahl et al (36) and recently validated by Fontaine et al (47), to obtain the double tg( lhb :hrGfpII/ fshb :DsRed2). In the double tg line, the two cell types expressing either fshb or lhb were clearly separated (Figure 1I-L).…”

“…Thus, we inferred that the dsred2 -positive cells produce Fsh hormone and can therefore be referred to as Fsh cells. We then crossed tg( fshb :DsRed2) fish with fish from the previously established tg( lhb :hrGfpII) line (36, 47) (Figure 1E-H) and examined the spatial distribution of Fsh and Lh cells. In agreement with prior studies in teleosts (7), in the double tg( lhb :hrGfpII/ fshb :DsRed2) medaka Fsh and Lh were expressed by distinct pituitary cells, thereby validating its use to simultaneously study Fsh and Lh cells (Figure 1I-L).…”