The small GTPase rab5 appears to be rate-limiting for the constitutive internalization of transferrin receptor and for fluid-phase endocytosis. However, it is unknown whether rab5 regulates receptors ...whose internalization is stimulated by the binding of ligand, and whether such receptors change the underlying rate of the endocytic pathways they utilize. As a model for ligand-stimulated endocytosis, we used transfected HEK293 cells expressing high levels of an epitope-tagged human beta 2-adrenergic receptor. Nearly all receptors were on the cell surface in the absence of agonist, but within ten minutes of agonist addition > 50% of receptors internalized and colocalized extensively with rab5. Hypertonic sucrose blocked beta 2-adrenergic receptor internalization, as well as that of transferrin receptor, suggesting a clathrin-mediated process. In contrast, an inhibitor of potocytosis had little effect upon beta 2-adrenergic receptor internalization, suggesting that this process did not require active caveolae. Consistent with this finding, caveolin was not detectable in the 12 beta 6 line, as assessed by western blotting with a polyclonal anti-caveolin antibody. Stimulated receptor internalization did not affect the rate or capacity of the constitutive endocytic pathway since there was no detectable increase in fluid-phase endocytosis after addition of beta-agonist, nor was there a significant change in the amount of surface transferrin receptor. Altogether, these data suggest that beta 2-adrenergic receptors internalize by a clathrin-mediated and rab5-regulated constitutive endocytic pathway. Further, agonist-stimulated receptor internalization has no detectable effect upon the function of this pathway.
B. R. Pitt, J. W. Ryan, A. Y. Chung, M. Woodford, D. B. Yeates and C. N. Gillis
Quantitative assessment of lung metabolic function is thought to provide
biochemical information reflecting integrity ...of the pulmonary
microcirculation. Although multiple indicator-dilution techniques are
useful in such pharmacokinetic studies, the need for fractionation and
subsequent processing of blood samples greatly prolongs data generation.
Accordingly, we designed and tested an in-line system which rapidly can
quantify single-pass disposition of photon-emitting substances in the
pulmonary circulation of intact animals. The nuclear detection system
consisted of a phoswich scintillation probe optically coupled to a
photomultiplier tube. Pulses were discriminated for height and shape and
counts recorded in a counter-timer, the output of which was interfaced with
a personal computer. A mixture of an intravascular reference substance
(99mTc-sulfur colloid) and an inhibitor of angiotensin-converting enzyme,
N-1(S)-carboxy-(4-OH-3-125I-phenyl) ethyl-L-alanyl-L-proline
(125I-CPAP), was injected as a bolus in the right heart of anesthetized
ventilated rabbits and arterial blood was diverted through a flow-cell
cuvette directly apposed to the phoswich detector. Single-pass extraction
of 125I-CPAP was 39 +/- 3% (mean +/- SE; n = 20) and was depressed in a
dose-dependent fashion by the addition of unlabeled CPAP (1-10
micrograms/kg) to the injection. These data indicate that we can now
quantify, in the intact animal, saturable binding of an inhibitor to
angiotensin-converting enzyme expressed on the surface of the pulmonary
microvascular endothelium. Furthermore, such data can be obtained rapidly.
This report summarizes the individual farm records of the members of the Southwestern Minnesota Farm Business Management Association for 2011. Whole-farm information and enterprise costs and returns ...are reported. The year-end analysis of the individual farms was performed by the Association fieldmen using the FINPACK software from the Center for Farm Financial Management. The individual analyses were summarized at the Department of Applied Economics using RankEm Central. In addition to the average of all farms, the averages for the high and low income groups are also presented. The tables are divided into four major groups: whole-farm information, crop enterprises, livestock enterprises, and summary information. The summary information includes whole farm historical trends and information by county, farm size, type of farm, debt to asset ratio, and operator age. Of the 114 farms in the Southwestern Association, the data for 107 farms are included in this report. The rest were omitted because the records were incomplete at the time that this report was prepared. In addition to this report, members receive an annual farm business analysis; on-farm instructional visits; end-of-year income tax planning and preparation; periodic meetings, tours, and seminars; a monthly newsletter; and other managerial and educational assistance. Each farmer pays an annual fee which covers the majority of the cost, with the balance defrayed by University of Minnesota Extension and research programs of the University of Minnesota.
PDF
