Flow Cytometric Analysis and Sorting of Live Male Rat Anterior Pituitary Cell Types by Forward Angle and Perpendicular Light Scatter*

Abstract

Light scatter patterns produced by living cells in the flow cytometer are known to provide useful information with regard to their size and internal structure. The purpose of this study was to determine if light scatter signals produced by live male rat anterior pituitary cells could be used as markers to aid in the identification and separation of different hormone-containing cell types. The typical light scatter pattern (4 X 10(5) cells/sample X 15 min) had three ridges in the forward angle light scatter (FALS) perpendicular light scatter (PLS) bivariate cell distribution. FALS signals could be correlated with the size of different cell types and PLS signals with their content of cytoplasmic secretory granules. Agranular cells dominated the low PLS ridge while moderately granulated PRL cells and heavily granulated GH cells dominated the medium and high PLS ridges, respectively. These light scatter patterns were reproducible both within and between different cell suspensions. Inclusion of dopamine in the pituitary gland dissociation medium, a treatment known to increase intracellular PRL contents of mammotrophs, increased the intensity of the PLS signals of a large population of cells, presumably PRL cells. Pituitary cells prepared from different aged male rats also showed changes in light scatter. Cell sorting on the basis of FALS-PLS signals established the relationship between cell type and light scatter pattern.

Triiodothyronine Reduces Growth Hormone Secretion and Pituitary Growth Hormone mRNA in the Chicken, in Vivo and in Vitro

“…The observed stimulatory effects of GHRF on GH mRNA on chicken pituitary cells is similar to that reported for rats (33,34), cattle (12), and sheep (35). These data are consistent with there being a common promoter mechanism for GHRF stimulating GH transcription and also GH release in mammals, birds, and their common ancestor-the divergence of mammals and birds occurring about 3 16 million years ago (40).…”

“…The flow cytometric immunofluorescence staining of anterior pituitary gland cells was performed as described by Hatfield and Hymer (35) with minor modifications (34). An aliquot of each pool of dispersed anterior pituitary cells was fixed with PBSbuffered 4% w/v paraformaldehyde for 30 min.…”

Prolactin Induction of Interleukin-2 Receptors on Rat Splenic Lymphocytes*

“…PRL (B6; 1 /ug/ml) was incubated with polyclonal antiserum (1:1000 final dilution) to rat PRL or GH (aGH) for 18 h at room temperature. Both antisera have been validated; their crossreactivity is 0.3% (15). The antibodytreated hormones were added to splenocytes, which were assayed for IL-2 responsiveness as described in Fig.…”

Flow cytometric analysis of functional anterior pituitary cells from female rats

“…It measures both cell size and granularity. Forward-or narrow-angle light scatter (FALS) is a good measure of cell size, whilst ninety degree or perpendicular light scatter (PLS) is related to cytoplasmic granulation and nuclear shape (Hatfield & Hymer, 1986a). By measuring angles of light scatter, a fluorescence-activated cell sorter (FACS) is capable of analysing and sorting cells on the basis of their size and granularity as well as fluor¬ escence.…”