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Biomedical subjects

M F Horster

Publications and source records attributed to M F Horster.

23 records · Page 2Linked to original sources

Assay of Na-K-ATPase in cultured nephron epithelia.

Microenzymatic methods have been utilized in the past to quantify the activity of Na-K-ATPase in tubular segments of the mammalian nephron. The assay reported here measures the precipitated inorganic phosphate liberated by the hydrolysis of gamma-32P-ATP. Activity data in single nephron segments of the cortical collecting tubule (CCT) confirm previous work; specifically, first data on enzyme activity in small cultured cell populations derived from CCT demonstrate that the Na-carrier enzyme can be quantified in nephron cell cultures.

Ammonia↗

Direct evaluation of fluorescence in single renal epithelial cells using a mitochondrial probe (DASPMI).

The study of distribution and quantitation of a fluorescent probe in living epithelia with the aid of an inverted microscope requires that individual cells can be analysed without optical interference from adjacent cells. This report describes the application of fluorescence microscopy and fluorometry to a recently developed in vitro culture system of renal epithelial cells. Epithelial cells derived from the mammalian renal cortical collecting tubule (CT) and the thick ascending loop of Henle (TAL) are cultivated as continuous monolayers in serum-free, hormone-supplemented media. A specific mitochondrial marker (DASPMI) is added to the medium and incorporated into the cytoplasm. The microscopic image reveals that the mitochondrial fluorescence distribution differs between CT and TAL cultures. The fluorometric quantitation shows a normally distributed histogram of medium-range intensity in TAL cell cultures while CT cultures exhibit a two-peak pattern of mitochondrial fluorescence distribution among epithelial cells.

Animals↗

Differential response to hormones of defined distal nephron epithelia in culture.

Defined cultures of rabbit kidney cortical collecting tubule (CCT) and cortical thick ascending limb of Henle's loop (CAL) were grown in monolayers from individual microdissected tubules and maintained for up to five passages, a maximum of 53 days. CCT cells contained cytochemically demonstrable vasopressin-stimulated adenylate cyclase, whereas CAL cells were characterized by the localization of Na+-K+-ATPase. [3H]thymidine labeling index decreased with time in primary cultures in the presence or absence of 3% serum. When added to unsupplemented serum-free media alone or in combinations, the growth factors dexamethasone, thyroxine, insulin, epidermal growth factor, and prolactin stimulated [3H]thymidine incorporation to different extents. CCT cells were maximally stimulated by addition of dexamethasone alone, whereas a combination of dexamethasone, thyroxine, insulin, and prolactin was most stimulatory for CAL cells. Addition of hormones concerned with renal ion and water transport to fully supplemented serum-free media inhibited [3H]thymidine labeling index: 1) vasopressin, isoproterenol, and dibutyryl cAMP were equally inhibitory in CCT and CAL cultures; 2) parathyroid hormone and prostaglandin E1 were more inhibitory in CAL cultures; and 3) aldosterone was particularly inhibitory in CCT cultures.

Aldosterone↗

Functional differentiation of the medullary collecting tubule: influence of vasopressin.

Medullary collecting tubules of rabbits were dissected from the outer zone at different stages of ontogenetic evolution and perfused in vitro. The hydraulic conductivity coefficient (Lp) was measured in the presence of either hypotonic perfusate and isotonic bath or isotonic perfusate and hypertonic bath. Basal Lp (cm s-1 atm-1 10(-7) was 85 +/- 34 (N = 17) during early functional differentiation of the outer medullary collecting tubule (e-OMCT), 36 +/- 6 (N = 8) in the intermediate state (i-OMCT), and 10 +/- 7 (N = 11) in the final, mature state of function (m-OMCT). Addition of supramaximal concentrations of arginine-vasopressin (AVP) to the bath increased Lp in i-OMCT (250 +/- 36) and m-OMCT (327 +/- 63) but did not activate the osmotic hydraulic conductance in e-OMCT (105 +/- 27). In 11 studies, OMCT were analyzed using isotonic solutions as the perfusate and bath medium. The spontaneous transtubular voltage (PD) was lumen positive, 1.71 +/- 0.3 in e-OMCT, lumen negative 2.43 +/- 0.3 in i-OMCT, and 6.1 +/- 0.4 in m-OMCT. AVP had no effect on PD in e- and i-OMCT but increased the PD significantly (P less than 0.025) to 9.2 +/- 0.6 in m-OMCT. The results indicate that both hydraulic conductivity coefficient and transtubular voltage in the medullary collecting tubule are dependent upon epithelial ontogeny and, particularly, display differential responses to the antidiuretic hormone. The data suggest that the differentiation of water and ion transport in the medullary collecting tubule may contribute to the increasing efficacy of the medullary countercurrent system.

Aging↗