Respiratory-related cortical potentials evoked by inspiratory occlusion in humans

It has long been recognized that humans can perceive respiratory loads. There have been several studies on the detection and psychophysical quantification of mechanical load perception. This investigation was designed to record cortical sensory neurogenic activity related to inspiratory mechanical loading in humans. Inspiration was periodically occluded in human subjects while the electroencephalographic (EEG) activity in the somatosensory region of the cerebral cortex was recorded. The onset of inspiratory mouth pressure (Pm) was used to initiate signal averaging of the EEG signals. Cortical evoked potentials elicited by inspiratory occlusions were observed when C3 and C alpha were referenced to CZ. This evoked potential was not observed with the control (unoccluded) breaths. There was considerable subject variability in the peak latencies that was related to the differences in the inspiratory drive, as measured by occlusion pressure (P0.1). The results of this study demonstrate that neurogenic activity can be recorded in the somatosensory region of the cortex that is related to inspiratory occlusions. The peak latencies are longer than analogous somatosensory evoked potentials elicited by stimulation of the hand and foot. It is hypothesized that a portion of this latency difference is related to the time required for the subject to generate sufficient inspiratory force to activate the afferents mediating the cortical response.     

Role of protein synthesis and proteases in production and inactivation of maturation-promoting activity during meiotic maturation of starfish oocytes

In starfish oocytes, activity of the maturation-promoting factor (MPF) and that of a major cAMP-independent protein kinase dropped at the time of meiotic cleavage, and rose again after the first but not the second meiotic cleavage. Protein synthesis was required before the first meiotic cleavage for both MPF and protein kinase activity to rise again after the first meiotic cleavage. Microinjection of either leupeptin or soybean trypsin inhibitor early enough prior to first polar body emission suppressed both the meiotic cleavage and the associated drop of MPF activity. Microinjection of leupeptin or soybean trypsin inhibitor during the 10-min period before the first meiotic cleavage also suppressed cytokinesis but did not prevent a decrease in MPF activity at the normal time of cytokinesis. The lysosomotropic inhibitor ammonia neither suppressed cytokinesis nor the drop of MPF activity at the time of first meiotic cleavage. Activity of neutral proteases sensitive to leupeptin and soybean trypsin inhibitor was demonstrated in oocyte homogenates prepared at the time of first meiotic cleavage. It is proposed that such proteases might be involved in degradation of protein kinase(s) and in the drop of MPF activity at the time of first meiotic cleavage.     

Reorganization of intermediate filament cytoskeleton in induced metanephric mesenchyme cells is independent of tubule morphogenesis

The metanephric mesenchyme becomes converted into epithelial tubules if cultured in transfilter contact with an inductor tissue. The expression of intermediate filaments (IFs), used as cell-type-specific markers has been studied in this model system for differentiation and organogenesis. In immunofluorescence microscopy of frozen sections, the undifferentiated cells of isolated metanephric mesenchymes uniformly showed IFs of vimentin type only. Also, when cultured as a monolayer, cells from the uninduced mesenchymes showed only vimentin filaments. In frozen sections of transfilter explants, epithelial tubules apparently negative for vimentin could be seen after 3 days in culture, but expression of cytokeratin could not be demonstrated in the developing tubules until the fourth day of culture. Sections of explants cultured further showed tubule cells with distinct fibrillar cytokeratin positivity. The appearance of cytokeratin in the explants was also demonstrated with immunoblotting experiments, using two different cytokeratin antibodies. Expression of IFs was further examined in monolayer cultures of metanephric mesenchymes which had been initially exposed to a short transfilter induction pulse. In these experiments, cytokeratin-positive cells could be demonstrated after a total of 4 days in culture. Double immunofluorescence experiments showed varying amounts of vimentin in the cytokeratin-positive cells: after 4 days in culture, most cytokeratin-positive cells still showed vimentin-positivity although often in a nonfibrillar form. During further culture, gradual disappearance of vimentin-specific fluorescence was observed in cytokeratin-positive cells. The results suggest that the vimentin-positive metanephric mesenchyme cells lose their fibrillar vimentin organization upon induction that leads to kidney tubule formation. This change may be essential for the transformation from an undifferentiated mesenchymal cell into a specialized epithelial cell. Cytokeratin filaments, regarded as a marker for epithelial cells, seem to appear simultaneously with or soon after the change in vimentin organization. These changes in IF expression also occur in monolayer cultures of mesenchyme cells initially exposed to a short transfilter induction pulse. This suggests that epithelial differentiation, as revealed by the emergence of cytokeratin positivity, may occur even in the absence of a clear morphological differentiation and three-dimensional organization of the cells.     

Phospholipid methyltransferase phosphorylation by intact hepatocytes: effect of glucagon

We have obtained a rabbit antiserum that specifically immunoprecipitates the 50K and 25K proteins of rat liver phospholipid methyltransferase. Exposure of intact rat hepatocytes preincubated with [32P]phosphate to glucagon induces a time-dependent phosphorylation of the 50K protein of phospholipid methyltransferase. The incorporation of 32P into the 50K protein was only on phosphoserine. These data support the concept that the activation of rat liver phospholipid methyltransferase by glucagon is mediated by phosphorylation of the enzyme.     

Sequential homologies between procarboxypeptidases A and B from porcine pancreas

Automated Edman degradation of monomeric procarboxypeptidases A and B from porcine pancreas shows that their N-terminal regions (from residue 1 to 34-37) present a high degree of sequential homology to each other as well as to other related procarboxypeptidases. Conformational predictions based on these sequences confirm their structural homology and indicate the probable existence of two beta-turns, one beta-chain and a long alpha-helix in them. On the other hand, tryptic peptide maps on a reverse-phase column indicate great sequential similarities (if not identity) between monomeric procarboxypeptidase A and the procarboxypeptidase A subunit isolated from its binary complex with proproteinase E.     

L-alanosine: a noncooperative substrate for Escherichia coli aspartate transcarbamylase

L-Alanosine, an antibiotic produced by Streptomyces alanosinicus, can be used by Escherichia coli aspartate transcarbamylase as a substrate instead of L-aspartate. The Michaelis constant of the catalytic subunit for this analogue is about 10 times higher than that for the physiological substrate, and the catalytic constant is about 30 times lower. The saturation curve of the native enzyme for L-alanosine indicates the lack of homotropic cooperative interactions between the catalytic sites for the utilization of this compound. It appears therefore that L-alanosine is unable to promote the allosteric transition. However, N-(phosphonoacetyl)-L-aspartate, a "bisubstrate analogue" of the physiological substrates, stimulates the reaction. This phenomenon is very similar to that reported by Foote and Lipscomb [Foote, J., & Lipscomb, W. N. (1981) J. Biol. Chem. 256, 11428-11433] concerning the reverse reaction using carbamylaspartate. The reaction is normally sensitive to the physiological effectors ATP and CTP. The significance of these results for the mechanism of the allosteric regulation is discussed.     

Fetuin modulates growth and differentiation of Ob17 preadipose cells in serum-free hormone-supplemented medium

A serum-free hormone-supplemented medium able to support the growth of rodent adipose precursor cells has been used to characterize additional components from serum required for the differentiation of preadipose Ob17 cells into adipose-like cells. Fetuin is shown to behave as a growth-promoting agent for these cells. In addition to growth hormone, triiodothyronine and a low-molecular weight component(s) also purified from serum, fetuin is required for the full expression of the differentiation program. Other serum proteins as well as other mitogenic factors are unable to substitute for fetuin. A possible role of fetuin in the development of adipose tissue is discussed.