Estrogen increases galanin immunoreactivity in hyperplastic prolactin-secreting cells in Fisher 344 rats.

Galanin is a widely distributed regulatory peptide which modulates the pituitary secretion of PRL and GH. Estrogen administration strongly stimulates galanin gene expression in the rat anterior pituitary. In adult female Fischer 344 rats, estrogen also induces hyperplasia of lactotropes. We used immunocytochemical analysis to assess the effects of estrogen on galanin-like immunoreactivity (Gal-IR) in the rat pituitary and hypothalamus during sc diethylstilbestrol (DES) implantation and after its removal at 30 days. In the anterior pituitary, DES implantation increased the portion of Gal-IR-containing cells from less than 2% in the control rats to 18.3% after 3 days of DES and 36% after 30 days. These changes paralleled the lactotrope hyperplasia exhibited in response to DES exposure. Ten and 30 days after removal of the DES capsules, the percentage of Gal-IR-containing cells in the anterior pituitary decreased to 6.3% and 1.5%, respectively. Colocalization studies revealed that Gal-IR-containing cells were predominantly lactotropes. Immunoelectron microscopy demonstrated that Gal-IR was concentrated in the Golgi region of these hyperplastic lactotropes and suggests that little of the synthesized galanin is secreted. The distribution of Gal-IR in the hypothalamus, median eminence, and neurohypophysis was unaffected by DES treatment. These data demonstrate that galanin is synthesized by hyperplastic pituitary lactotropes of Fischer 344 rats and that peptide accumulation is dependent on the presence of circulating estrogens. In contrast, neuronal galanin synthesis in the hypothalamus does not appear to be regulated by estrogen.

Lead staining in the Golgi complex.

Lead ions at similar concentrations to those used for Gomori type phosphatase localization stain some parts of the vacuolar system, particularly compartments of the Golgi complex (GC) and isolation envelopes (im) in a characteristic way in both vertebrates and invertebrates. After fixation in 2.5% glutaraldehyde, lead citrate in acetate or aspartate buffer (pH 5.5-7.2) leaves the contents of GC cisternal compartments with a fine particulate stippling. In the fat body of Calpodes ethlius and in mouse pancreas the staining is faint but definite without further enhancement of contrast, although it is easily overlooked after section staining. The distribution of lead stain differs from that of the lead phosphate precipitated after Gomori type acid phosphatase reactions. Whereas lead stain may be in all GC and im compartments, acid phosphatase is restricted to the innermost saccules and nearby vacuoles. The compartment specific staining by led also differs from the generalized staining in all compartments given by uranyl. Thus the contents of luminal membrane surfaces of some parts of the vacuolar system can be characterized by their ability to bind lead. In cells where protein synthesis has been blocked by cycloheximide, secretory vesicles are absent and the RER and GC from the generalized staining in all compartments given by uranyl. Thus the contents of luminal membrane surfaces of some parts of the vacuolar system can be characterized by their ability to bind lead. In cells where protein synthesis has been blocked by cycloheximide, secretory vesicles are absent and the RER and GC from the generalized staining in all compartments given by uranyl. Thus the contents of luminal membrane surfaces of some parts of the vacuolar system can be characterized by their ability to bind lead. In cells where protein synthesis has been blocked by cycloheximide, secretory vesicles are absent and the RER and GC cisternae are devoid of uranyl stainable material. However, lead staining and acid phosphatase activity in the GC continue. We presume that they mark the environment within these cisternae rather than the proteins passing through them. This environment is itself not static. Several observations suggest that the function of cisternae that is detectable by lead staining is temporally discontinuous and related to a stage of maturation or development. Only early stage ims stain: the staining ceases by the beginning of autophagy after hydrolytic enzymes are presumed to have been added. Condensing vacuoles cease to stain as the central core crystallizes out. Stain may be absent from one or two GC saccules at any position in the stack as though the phase of lead staining (or lack or it) can move progressively through the system. We conclude that in studies characterizing components of the vacuolar system it is necessary to separate those that mark transient occupants of a compartment from those that mark the compartment itself. Both may vary temporally independently from one another.

High resolution microanalysis for phosphorus in Golgi complex beads of insect fat body tissue by electron spectroscopic imaging.

Golgi complex beads are 10 nm particles arranged in rings on the smooth forming face of the Golgi complex that stain specifically with bismuth in arthropod cells. In vitro experiments with biological molecules spotted on to cellulose acetate strips indicated that bismuth bound to the beads through phosphate groups. We could detect a weak phosphorus signal from the beads using a new technique called electron spectroscopic imaging that is capable of very high spatial resolution (0.3-0.5 nm) and sensitivity (50 atoms of phosphorus). Detection was not obscured by tissue staining with bismuth or uranyl acetate of by using an inorganic buffer (Na cacodylate). Localization of phosphorus was greatly improved by using colour-enhanced computer pictures of the electron spectroscopic images and quantitating the images. The results indicate that the phosphorus content of the beads is large enough to account for their bismuth reactivity.

Tannic acid-metal salt sequences for light and electron microscopic localization of complex carbohydrates.

Use of tannic acid (TA), in sequence with ferric chloride, uranyl acetate or gold chloride resulted in staining of selective but sometimes different sites in paraffin sections. TA-uranyl acetate of TA-ferric chloride stained sites rich in complex carbohydrates, wherease TA-gold chloride stained the collagen of various connective tissues different shades of red-purple to gray-black. Applied to epoxy-embedded thin sections of tissues fixed with glutaraldehyde and not post-osmicated, TA-uranyl acetate and TA-ferric chloride imparted density to subcellular sites known to contain a high concentration of mucosubstances, such as secretory granules and cisternae of the Golgi complex of certain cells. TA-gold chloride proved unsatisfactory for ultracytochemistry because of its tendency to form globular precipitates on thin sections. The effect of blockage procedures at the light microscopic level indicated that vicinal glycols are not required for binding of TA to tissue sites. Electrostatic forces were shown to be of minimal significance, whereas hydrogen bonding appeared to play a part in both TA-tissue and TA-metal binding mechanisms.