Ronald Portanova

Isolated Pituitary Cells: CRF-Like Activity of Neurohypophysial and Related Polypeptides

Summary Neurohypophysial and structurally related compounds elicit secretion of ACTH from suspensions of isolated pituitary cells. However, the maximum secretion of ACTH in response to the polypeptides is significantly less than that for an extract of rat hypothalamic median eminence (HME) tissue. When added in combination with appropriate doses of HME extract, argvasopressin inhibits ACTH secretion. These findings suggest that the neurohypophysial polypeptides may be partial agonists of corticotropin secretion. The authors are grateful to Beth Wiblin-Portanova and James Larry Simmons for their technical assistance.

A Trypsin Technic for the Preparation of Isolated Rat Anterior Pituitary Cells

Summary Isolated rat anterior pituitary cells have been prepared by mechanical agitation of quartered glands in the presence of trypsin. The cells appear morphologically and functionally intact and contain large amounts of ACTH. The medium separated from cells incubated at 37° for 20 min contains small but significant amounts of ACTH. Acetic acid extracts of rat HME or cerebral cortical tissue and arginine vasopressin increase the ACTH content of the medium. The authors acknowledge the technical assistance of Mary Vegh and Beth Wiblin.

Corticosterone suppression of ACTH secretion: Actinomycin D sensitive and insensitive components of the response

Summary Corticosterone suppresses ACTH secretion by isolated pituitary cells. Actinomycin D (Act D) added simultaneously with, or slightly prior to, the steroid prevents the suppressive action on cells from adrenalectomized animals but not on cells from intact animals. A working hypothesis encompassing these findings is proposed: corticosterone induces the formation of a cellular factor with which it subsequently interacts to suppress ACTH secretion. The formation of the factor is sensitive to Act D, the subsequent interaction with corticosterone is not. The factor is present in the pituitary cells of intact animals owing to their previous in vivo exposure to endogenous steroid; it is absent from the pituitary cells of adrenalectomized animals.

Effect of glucocorticoid levels in vivo on growth hormone biosynthesis.

The rate of growth hormone (GH) biosynthesis in pituitary cells prepared from rats with different histories of glucocorticoid exposure was analyzed by a dual-labeling acrylamide-gel technique. Glucocorticoid deficiency, produced by adrenalectomy, reduces GH synthesis by approximately 50%. GH synthesis adrenalectomized rats treated with either natural or synthetic glucocorticoids, is restored to normal or above normal levels. Acute exposure (1 h) of cells to corticosterone in vitro does not produce a significant increase in GH synthesis. Adrenalectomized rats treated with steroid for one day show a small but detectable increase in GH synthesis, while treatment for 3--6 days results in progressively larger stimulatory effects. All of these results are similar to previous findings regarding glucocorticoid stimulation of GH synthesis in vitro.

Release of ACTH from isolated pituitary cells: an energy dependent process.

Summary The influence of various maneuvers designed to alter energy metabolism on spontaneous and induced release of ACTH from isolated pituitary cells was studied. Spontaneous release was increased by cold (2°) but not significantly changed by anoxia, glucose removal, DNP, iodoacetate, oligomycin, or antimycin A. On the other hand, induced release of ACTH was inhibited by all of the maneuvers tested. These findings indicate that induced release of ACTH from isolated pituitary cells is an energy dependent process. The author is grateful to Dr. George Sayers for his invaluable advice during the conduct of these experiments and preparation of the manuscript and to Beth Portanova and Marshall Clark for their technical assistance.

Fine Structure of Trypsin-Dissociated Cells of the Rat Anterior Pituitary Gland

Summary Trypsin-dissociated cells from the rat anterior pituitary appeared essentially intact and undamaged in the electron microscope. Suspensions contained five cell types which correspond closely to those seen in situ: thyrotroph (with cytoplasmic granules 1500 Å approximate maximal diam), corticotroph (2000 Å), gonadotroph (2000 Å), somatotroph (4000 Å), and mammotroph (7500 Å).

Corticosterone induction of pituitary hyporesponsiveness to CRF: Divergent effects of puromycin and cycloheximide

Abstract Isolated pituitary cells prepared from adrenalectomized rats secrete ACTH in response to corticotrophin releasing factor (CRF) contained in extracts of rat hypothalamic-median eminence tissue (HME-CRF). Neither puromycin nor cycloheximide, at doses which inhibit protein synthesis by up to 86%, significantly affect HME-CRF stimulated ACTH secretion. When pituitary cells are exposed to corticosterone, after a lag of approximately 30 min they become hyporesponsive to stimulation by HME-CRF. Graded doses of puromycin inhibit this action of the steroid hormone to the same degree that they inhibit protein synthesis. This quantitative correlation suggests that protein synthesis is an intermediate step in the mechanism of corticosterone action on the pituitary. On the other hand, cycloheximide does not block corticosterone action, even at concentrations of cycloheximide which inhibit protein synthesis by 90%. Although the basis for these divergent effects is not yet clear, several important implications are discussed.

The Role of Cyclic AMP in CRF-Induced ACTH Secretion

Summary ACTH secretion by isolated pituitary cells is stimulated both by HME-CRF and DBC, and when given in combination, the two secretagogues interact synergistically. Although the mechanism of this interaction is unknown, the potentiating effect of DBC is displayed without an apparent time lag and persists after removal of the cyclic nucleotide. Corticosterone inhibits ACTH secretion induced by HME-CRF and DBC, acting alone or in combination. The implications of these findings are discussed. The authors are grateful to Beth Wiblin Portanova and James Roe for their expert technical assistance.