Albumin phylogeny for clawed frogs (Xenopus).

Comparisons of albumin indicate that the frogs commonly used by North American molecular and developmental biologists under the name of Xenopus muelleri belong to another species, X. borealis. Phylogenetic analysis of the albumin data reveals two major groups of Xenopus species, one containing only X. tropicalis and the other, called the X. laevis grou, containing the remaining species of the genus. The phylogenetic tree, in conjunction with evidence from chromosomes and DNA content, leads to the hypothesis that total genome duplication occurred in the common ancestor of the X. laevis group.

Alterations in electrophysiology of cultured BALB/cfC3H mouse mammary epithelium associated with neoplastic transformation.

Mammary tumor virus and neoplastic transformation affect prolactin-modulated electrophysiology of mouse mammary epithelium in vitro. Transepithelial resistances of both prelactating and neoplastic BALB/cfC3H mouse mammary epithelial cell cultures are lower than those seen in BALB/c prelactating cell cultures. Unlike the response to prolactin in BALB/c cultures, there are no effects of this hormone on transepithelial potential difference or resistance of BALB/cfC3H prelactating or neoplastic cultures. However, prolactin significantly increases short-circuit current in both BALB/c and BALB/cfC3H prelactating cell cultures and in BALB/cfC3H neoplastic cell cultures. The response of tumor cell cultures suggests that pregnancy-independent mouse mammary epithelium may not be entirely refractory to prolactin.

Transepithelial electrophysiology of cultured mouse mammary epithelium: sensitivity to prolactins.

Several prolactins and growth hormones affect cultured BALB/c mouse mammary epithelial electrophysiology at doses of 10 micrograms/ml. Ovine prolactin is effective at doses as low as 1 ng/ml, levels that are three to four orders of magnitude less than effective amounts of rat growth hormone. Because growth hormone preparations may have approximately 1% prolactin contamination, its effects are probably due to prolactin contamination. The maximum effect of ovine prolactin on short-circuit current and on resistance occurs at doses differing by two orders of magnitude, indicating that these two effects of prolactin may be partially separable. Therefore, prolactin may have effects on passive permeability properties in addition to its documented effects on active transport.

Prolactin effects on ion transport across cultured mouse mammary epithelium.

Prolactin is a known osmoregulatory hormone in lower vertebrates, and recent evidence indicates that this hormone modulates ionic concentrations in milk. In an ultrastructurally and biochemically differentiated primary cell culture system in which mouse mammary epithelium is maintained on floating collagen gels, prolactin causes an increase in short-circuit current (Isc) of monolayers of cells derived from midpregnant (24.6 to 48.0 microA . cm-2) and lactating (10.4 to 16.1 microA . cm-2) glands. Transepithelial potential differences (basal side ground) average about -12 mV and are similar to those seen in vivo. Prelactating mammary epithelial cell cultures have transepithelial resistances ranging from 374 omega . cm2 (prolactin present) to 507 omega . cm2 (prolactin absent), and lactating cell cultures have resistances averaging almost 1,000 omega . cm2. Prolactin effects require at most one day of culture maintenance in prolactin-containing medium, and the effects are not due to known contamination of prolactin preparations with arginine vasopressin or growth hormone. Medium concentrations of prolactin as low as 1 ng/ml can elicit these effects. In prelactating cell cultures not treated with prolactin, the Isc is equal to the rate of sodium absorption. Prolactin increases sodium absorption fourfold but increases Isc only twofold. Clearly, prolactin induces other active transport; neither potassium nor chloride movements can account for this additional transport. Resistance values, current-voltage plots, and permeability coefficients indicate that in vitro mammary epithelium is a moderately "tight" tissue. Comparisons with intact glands indicate that in vitro mammary epithelium closely resembles its in vivo counterpart. Floating collagen gel cultures appear suitable for elucidating transport properties in cellularly heterogeneous and structurally complex mammalian tissues.

Mouse mammary epithelial cells on floating collagen gels: transepithelial ion transport and effects of prolactin.

Epithelial cells dissociated from midpregnant BALB/c mouse mammary glands were cultured for as long as 20 days as confluent monolayers on floating collagen gels. Detached gels bearing monolayers were placed in lucite Ussing chàmbers for measurement of transepithelial potential difference (PD), short-circuit current (I(sc)), resistance (R), and unidirectional fluxes of Na(+) and Cl(-) during short-circuit current conditions (PD = 0). With Hanks' solution bathing both sides of cultures maintained with insulin and cortisol, PD = -12.8 mV (serosal side ground), I(sc) = 24.6 muA/cm(2), and R = 507 omega.cm(2). Net absorption of Na(+) equaled I(sc), and there was no net Cl(-) transport. PD and I(sc) were reduced 50% by mucosal addition of 10 muM amiloride and to zero by metabolic inhibition with nitrogen gas or by serosal addition of 0.1 mM ouabain. In similar cultures supplemented with prolactin, PD and I(sc) increased to -15.8 mV and 48.0 muA/cm(2), respectively, and R decreased to 374 omega.cm(2). Inhibitor effects were similar to those seen in prolactin-free cultures. Prolactin exposure resulted in a 3-fold increase in net absorption of Na(+). Na(+) absorption was not equivalent to I(sc), and there was little Cl(-) absorption; therefore, prolactin induced active transport of other, as yet unidentified, ions. These effects of prolactin require at least 3 days to occur and cannot be attributed to the known contamination with neurohypophysial hormones. The prolactin-induced increase in Na(+) absorption parallels its Na(+)-retaining ability in lower vertebrates and could be part of the mechanism that keeps milk Na(+) concentration low in intact glands.

Fetal rabbit gastric epithelial cells cultured on floating collagen gels.

Cells were isolated from approximately 30 d fetal rabbit stomachs and cultured on floating collagen gels. Electron microscopy showed monolayers in which only one cell type persisted. These columnar cells were joined at apical borders by tight junctions and contained an extensive endoplasmic reticular network with an occasional intracellular canaliculus. They also occasionally contained what appeared to be secretory granules (mucus?), and therefore had some characteristics of all the cell types of the intact fetal stomachs, which showed oxyntic, mucous, and undifferentiated cells. In Ussing chambers with Ringer's solution on both sides, cultures developed transepithelial potential (potential difference [PD], mV, mucosa ground) = 13, resistance (resistance [R], omega - cm2) - 285, and short-circuit current (Isc, microA/cm3) = 45 (n = 7), clearly indicating that cellular polarity and junctional integrity were maintained. These transport parameters were somewhat different for intact fetal stomachs (PD = 20, R = 70, and Isc = 220 [n = 4]), which may be due to extensive folding of intact fetal stomachs or the presence of only one cell type in culture, or both. Although gastric stimulants histamine, dibutyryl cycle AMP (dbcAMP), and isobutylmethylxanthine (IMX) (a phosphodiesterase inhibitor) did not elicit H+ secretion or electrophysiological changes in monolayers or intact stomachs, 10-4 M apical amiloride caused a decrease in Isc in cultured monolayers(27%) and intact stomachs (50%). Thus, Na+ transport seems to be a significant fraction of ion transport in both preparations. This culture system may allow the study of oxyntic cell differentiation and the development of H+, Na+, and Cl- transport in the gastric musoca.