Melanocortin‐1 receptor structure and functional regulation

The melanogenic actions of the melanocortins are mediated by the melanocortin‐1 receptor (MC1R). MC1R is a member of the G‐protein‐coupled receptors (GPCR) superfamily expressed in cutaneous and hair follicle melanocytes. Activation of MC1R by adrenocorticotrophin or α‐melanocyte stimulating hormone is positively coupled to the cAMP signaling pathway and leads to a stimulation of melanogenesis and a switch from the synthesis of pheomelanins to the production of eumelanic pigments. The functional behavior of the MC1R agrees with emerging concepts in GPCR signaling including dimerization, coupling to more than one signaling pathway and a high agonist‐independent constitutive activity accounting for inverse agonism phenomena. In addition, MC1R displays unique properties such as an unusually high number of natural variants often associated with clearly visible phenotypes and the occurrence of endogenous peptide antagonists. Therefore MC1R is an ideal model to study GPCR function. Here we review our current knowledge of MC1R structure and function, with emphasis on information gathered from the analysis of natural variants. We also discuss recent data on the regulation of MC1R function by paracrine and endocrine factors and by external stimuli such as ultraviolet light.     

A general,fast, and sensitive micromethod for DNA determination: Application to rat and mouse liver,rat hepatoma,human leukocytes,chicken fibroblasts,and yeast cells

A fluorometric method using 3,5-diaminobenzoic acid for DNA determination in tissues, cultured cells, nucleated blood cells, and yeast cells is described. The method is general, simple, and rapid, and does not require prior DNA extraction, since tissue is directly solubilized in Triton X-100 and ammonia. The procedure is highly sensitive, and is able to measure rather accurately as little as 10 ng of DNA. It is applicable to all types of DNA structure. The DNA content determined in various tissues and cells was: 2.50 mg/g fresh rat liver, 3.32 mg/g rat diethylnitrosamine-induced hepatoma, 2.49 mg/g fresh mouse liver, 8.76 μg/10⁶ human leukocytes, 3.37 μg/10⁶ chicken fibroblasts, 2.97 μg/10⁸ haploid yeast cells, and 2.84 μg/10⁸ haploid yeast protoplasts.     

ATP stimulates inositol phosphates accumulation and calcium mobilization in a primary culture of rat aortic myocytes

The effects of extracellular ATP on phosphoinositide metabolism and intracellular Ca2+ concentration were studied in a primary culture of rat aortic myocytes. ATP increases the level of inositol phosphates, the putative second messenger for Ca2+ mobilization. No saturation of inositol phosphates accumulation is obtained (up to 10(-2) M ATP). Under the same conditions, ATP rapidly mobilizes intracellular Ca2+ in fura-2 loaded myocytes. The mobilization of intracellular Ca2+ is dose-dependent (maximal at 10(-4) M ATP), and is not affected by addition of EGTA. It is concluded that the receptors mediating the cytosolic increase of Ca2+ are of the P2-purinoceptor subtype. The physiological functions of these receptors are not presently known.     

The receptor for human sex steroid binding protein (SBP) is expressed on membranes of neoplastic endometrium.

Sex steroid binding protein (SBP) receptor was detected on cell membranes obtained from human endometrium adenocarcinoma. The binding of SBP was proved to be highly specific, saturable, and at high affinity. It was, additionally, shown to occur at two sites at different affinities, as previously described for other human tissues. SBP was, therefore, demonstrated to recognized a specific receptor on endometrium adenocarcinoma membranes. The effect of steroid hormones on SBP-receptor interaction was also evaluated. Both dihydrotestosterone and estradiol were shown to inhibit the binding of SBP to its specific receptor on neoplastic membranes. Testosterone at a dose of 10(-9) M was shown not to interfere to a significant extent with SBP-receptor binding. The sensitivity for estradiol we had previously observed in normal premenopausal endometrium was completely lost in postmenopausal neoplastic tissue. These observations suggest that the SBP-membrane recognition system is still present in neoplastic postmenopausal endometrium, but it has been modified either by the postmenopausal endogenous milieu or by the neoplastic transformation.     

Histophysiological studies on the corpus allatum of Leucophaea maderae

Summary The structural differences between active and inactive corpora allata, visible under the light microscope, become more pronounced under the electron microscope. Aside from differences in cellular and nuclear diameters, and nuclearcytoplasmic ratios, there are qualitative characteristics in ultrastructural organization.The cytoplasm of active corpus allatum cells contains numerous sinuous mitochondria, distinct Golgi elements, ergastoplasmic units with a tendency to form whorls, agranular cytomembranes, and free ribosomes. Pleomorphic inclusion bodies resembling lysosomes are more or less numerous. The plump, ovoid nuclei frequently show two prominent nucleoli whose components may form a meshwork harboring chromatin.The marked reduction in the amount of cytoplasm occurring during the organ's return to inactivity is accompanied by a decrease in the number, and a change in the appearance, of some cytoplasmic organelles. The mitochondria tend to be smaller, and the ergastoplasm is reduced to scattered wisps of ribosome-studded membranes. Nuclei of inactive cells have smaller diameters than those of active ones.In all stages of activity, cell boundaries are clearly visible. As a result, the corpus allatum cell can now be characterized as a discrete unit of epithelioid character and rather complex shape. The plasma membrane may become folded when the cellular content shrinks to the inactive level. Aside from changing outlines, all corpus allatum cells have long, gradually thinning processes. These penetrate deeply into the parenchyma where they interlock with those of other cells; many processes eventually seem to reach the surface of the gland where the secretory products are released into the hemolymph. These have to pass through an acellular connective tissue layer that shares tinctorial and ultrastructural properties with those of a boundary (or basement) membrane.This stromal element forms a sheath and branching processes that extend into and permeate the parenchyma. It seems to represent a system of channels, not only for the release of secretory and other cellular products, but for the entry of nutrients and perhaps chemical messenger substances.Neurosecretory material can be observed in the form of structurally distinctive elements, i.e., as electron-opaque granular inclusions within axon terminals that become contiguous with corpus allatum cells.No definite statement can be made on the basis of the present study about the nature of the corpus allatum hormone or hormones, except that the ultrastructural criteria indicative of proteinaceous secretion, such as the appearance of secretory granules in spatial relation with Golgi elements, seem to be missing in the corpora allata of Leucophaea.Supported by Research Grants A-3984 and B-2145 from the U.S.P.H.S.     

Interactions between the soilborne root pathogenPhytophthora nicotianae var.parasitica and the arbuscular mycorrhizal fungusGlomus mosseae in tomato plants

In order to study the influence of Arbuscular Mycorrhiza (AM) on the development of root rot infection, tomato plants were raised with or withoutGlomus mosseae and/orPhytophthora nicotianae var.parasitica in a sand culture system. All plants were fed with a nutrient solution containing one of two phosphorus (P) levels, 32µM (I P) or 96µM (II P), to test the consequence of enhanced P nutrition by the AM fungus on disease dynamics. Mycorrhizal plants had a similar development to that of control plants. Treatment withPhytophthora nicotianae var.parasitica resulted in a visible reduction in plant weight and in a widespread root necrosis in plants without mycorrhiza. The presence of the AM fungus decreased both weight reduction and root necrosis. The percentage reduction of adventitious root necrosis and of necrotic root apices ranged between 63 and 89% The enhancement of P nutrition increased plant development, but did not appreciably decrease disease spread. In our system, mycorrhiza increased plant resistance toP. nicotianae var.parasitica infection. Although a contribution of P nutrition by mycorrhiza cannot be excluded, other mechanisms appear to play a crucial role.     

Effect of osmotic stress on the growth of epicotyls of Cicer arietinum in relation to changes in the autolytic process and glycanhydrolytic cell wall enzymes

Simulation of drought by polyethylene glycol (PEG) inhibited elongation of epicotyls of Cicer arietinum L. cv. Castellana but had no effect on growth capacity since growth was restored once the inhibitory condition had been removed. The amount of proteins in the cell wall was correlated with the elongation of the epicotyls and decreased when elongation was inhibited. PEG-induced inhibition of elongation had different effects on the various glycanhydrolytic cell wall enzymes. Only α-galactosidase (EC 3. 2. 1. 22) seemed related to the lack of elongation, increasing its activity when elongation was inhibited. The β-galactosidase (EC 3. 2. 1. 23) and β-glucosidase (EC 3. 2. 1. 21) studied did not show changes in their specific activities during the inhibition of elongation. β-Galactosidase is responsible for the autolytic process in Cicer arietinum . This enzyme hydrolyzes specified linkages in the cell wall, releasing sugar constituents. Our present results show that β-galactosidase is not directly related with elongation because no changes could be observed during inhibition of elongation. The autolytic process is related with chemical processes taking place in the cell wall and preceding elongation of the epicotyls, i. e. the loosening process. Cell wall loosening is necessary for elongation to take place but elongation does not necessarily follow loosening if the osmotic conditions are unfavorable     

Neurosecretion. XIII. The ultrastructure of the corpus cardiacum of the insect Leucophaea maderae

Summary The corpora cardiaca of Leucophaea maderae contain two classes of intrinsic elements, parenchymal and interstitial cells. The parenchymal cells produce a secretory material first visible in the Golgi zones of the perikarya in the form of distinct electron-opaque granules. These undergo changes (gradual loss of electron-density, emergence of internal structure) as they accumulate in cellular processes.The parenchymal cells are best classified as neuroglandular elements since, in addition to secretory inclusions, they possess characteristics of ganglion cells such as axonlike processes, neurotubules, and sheaths. These covers are provided by branches of the second type of intrinsic elements, the interstitial cells. They are non-glandular structures of considerable morphological complexity. In the manner of glial elements, they permeate the entire organ and encapsulate not only the perikarya of parenchymal cells but cellular processes as well.The cytoplasmic processes include a) relatively short ones belonging to parenchymal cells, and b) long axons whose cell bodies lie within the central nervous system. Many of the latter contain electron-opaque granules of the kind found in electron-micrographs of typical neurosecretory cells. These extrinsic granules represent the second category of secretory products stored within the corpora cardiaca. By comparison with the product of the intrinsic gland cells, the neurosecretory granules seem to be fairly stable. Neither type seems to pass through the connective tissue sheath of the corpus cardiacum in the form of discrete granular entities.This sheath, which sends branches into the interior of the corpora cardiaca, has the properties of a basement membrane. It represents a pathway for the exchange of substances between the cellular components of the corpus cardiacum and the surrounding hemocoele.The dual character of the corpus cardiacum, namely that of a storage and release center for extrinsic neurosecretory substances and of an endocrine organ in its own right, is herewith established beyond doubt. The number of secretory products discernible on the basis of their morphology and localization (two size categories of extrinsic and one intrinsic type of granules) does not match the variety of physiologically active principles known at present. The assignment of specific functions to discrete morphological elements must await further studies.Supported by Research Grants A-3984 and B-2145 from the U.S.P.H.S.