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Biomedical subjects

R E Garfield

Publications and source records attributed to R E Garfield.

At least 163 records · Page 9Linked to original sources
PubMed1984

Control of myometrial function in preterm versus term labor.

In this review I have attempted to briefly describe myogenic, neurogenic, and hormonal mechanisms which control myometrial function during term and preterm labor. The synthesis of myometrial gap junctions and receptors in response to hormonal changes appears to be a key element in the gradual evolution of contractility leading to labor. I suggest that the development of myometrial gap junctions is physiologically regulated and can be manipulated pharmacologically. Thus, this system provides a means of regulating myogenic control of smooth muscle. I have proposed a model, or working hypothesis, which may explain the indirect and direct interactions of various substances involved in the control of synthetic processes and stimulating or inhibiting myometrial contractility. The model predicts that term or preterm labor can be initiated at various steps. A more complete understanding of these interactions is necessary for a rational approach to the control of term or preterm labor.

Electrophysiology↗
PubMed1984

Smooth muscle cell herniation in the contracted arterial wall of spontaneously hypertensive and normotensive rats.

Structural changes of blood vessels from hypertensive animals during contraction were studied using light and transmission microscopy. In the arteries, maximally contracted with norepinephrine, vacuoles were found in the media of three categories of mesenteric arteries from two age groups (10-12, 28-30 weeks) of spontaneously hypertensive (SHR) and age-matched normotensive Wistar-Kyoto (WKY) rats. These vacuoles were related to contraction of the smooth muscle cells involving shape change, as they were absent in relaxed vessel walls, and also in muscles that underwent isometric contraction. In the SHR, the number of vacuoles/1,000 microns 2 of media was positively correlated with the degree of contraction of the vessels. Electron microscopic examination of these vacuoles showed that they were the result of herniation (or evagination) of smooth muscle cells into the intercellular space as well as into the neighbouring cells. When the number of herniae per unit cross-sectional area of the contracted media was used as the basis for comparison between SHR and WKY, we found no difference between the two strains of rats at 10-12 weeks of age. At 28-30 weeks, however, the number of herniae was significantly lower in the SHR as compared with age-matched WKY and younger SHR, suggesting that functional changes, that functional changes, probably related to the physical property of the plasma membrane in the smooth muscle cells, have taken place in older SHR.

Animals↗
PubMed1983

Effect of estradiol-17 beta and prostaglandins on rat myometrial gap junctions.

The effects of estradiol-17 beta and indomethacin on myometrial gap junction development, plasma estradiol levels and uterine PGF2 alpha content were evaluated in immature and/or ovariectomized, mature rats. High doses of estradiol stimulated the development of gap junctions in the myometrium of animals from both groups. Concomitant injections of estradiol and indomethacin to ovariectomized rats potentiated the estradiol stimulation of gap junctions. Plasma estradiol levels were lower in ovariectomized rats treated with both estradiol and indomethacin than in animals treated with estradiol alone. Indomethacin also enhanced the uptake and retention of 3H-estradiol into uterine tissues. Uterine PGF2 alpha content of ovarectomized rats was stimulated with the initial injection of estradiol but thereafter, the PGF2 alpha content declined with repeated injections to values lower than that observed in controls. Prostaglandin F2 alpha content in tissues from rats treated with estradiol plus indomethacin were also higher than that observed in rats treated with indomethacin alone, however, the values obtained in both groups were significantly lower compared to those from control animals. These results are consistent with the hypothesis that steroid hormones and prostaglandins regulate myometrial gap junction formation. Regulation of myometrial gap junctions by prostaglandins is discussed with respect to a down regulation of the steroid-receptor mechanism and effects on cyclo-oxygenase or lipoxygenase products.

Animals↗
PubMed1983

Studies on canine gastric antrum smooth muscle: preparation and characterization of a plasma membrane enriched fraction.

A method is described for preparation of large amounts of a plasma membrane (PM) enriched fraction from the smooth muscle of dog antrum. It consists of preparing microsomes, treating them with ATP + EGTA + Mg, centrifuging in 30% sucrose and then centrifuging the resulting supernatant in 15% sucrose to yield the plasma membrane enriched fraction P6. The subcellular fractions obtained at various steps during purification were characterized by: 5'-nucleotidase and phosphodiesterase I as plasma membrane markers; cytochrome c oxidase as an inner mitochondrial marker; NADPH-cytochrome c reductase as a putative endoplasmic reticulum marker; electron microscopy; polyacrylamide sodium dodecyl sulfate slab gel electrophoresis. The distribution of ATP-dependent and independent Ca uptake in presence and absence of azide and the effect of 5 mM oxalate or 25 mM phosphate on this uptake was also examined. The fraction P6 consists of mostly smooth surface vesicles 164.3 +/- 7.2 nm in diameter, has an exclusion volume of 9.7 microL/mg for [3H]inulin and 11.1 microL/mg for [3H]sucrose. P6 is maximally enriched in the ATP-dependent azide-insensitive Ca-uptake capacity and as compared with the postnuclear supernatant (S1) it shows a very small percent stimulation by oxalate and phosphate. The ATP-dependent Ca uptake by the P6 fraction occurs optimally at pH 7.0-7.4 and is much larger than the ATP-independent Ca uptake. At pH 7.1, the ATP-dependent Ca uptake occurs with a Km of 0.27 microM and a Hill coefficient greater than 2 for Ca2+. Half maximum binding of Ca2+ occurred at 300 microM Ca2+. Ca ionophores A23187 and ionomycin inhibited the ATP-dependent Ca uptake, and if added after the uptake, these caused a release of the accumulated Ca2+. From these and other data it is concluded that this PM preparation contains a Ca transport system which can lead to formation of greater than 1000-fold Ca2+ concentration gradient across the vesicle membrane in 1 min when extravesicular Ca2+ concentration is 0.3 microM. Thus this preparation is an extremely useful material for studying the mechanism of action of the Ca pump in smooth muscle plasma membrane.

Adenosine Triphosphate↗
PubMed1983

Morphometric study of structural changes in the mesenteric blood vessels of spontaneously hypertensive rats.

Structural changes of three categories of mesenteric arteries (representing elastic, muscular and arteriolar vessels) from 10- to 12-week-old and 28-week-old spontaneously hypertensive rats (SHR) were studied morphometrically at the light microscope level, and the results compared with age-matched Wistar-Kyoto normotensive rats. In 10- to 12-week-old SHR, hypertrophy of the vessel wall occurred only in the muscular and arteriolar vessels. At 28 weeks, further thickening of the vessel wall occurred in the muscular and arteriolar vessels, and the superior mesenteric artery (elastic vessel) was also thickened in the SHR. There was no evidence that the wall of the relaxed hypertrophied vessels encroached upon the lumen of the vessel. The structural basis for the increase in the vessel wall thickness varied with vessel type. In the superior mesenteric artery, increase in the media at 28 weeks of age would be consistent with hypertrophy of the smooth muscle cells. In the large muscular arteries, at 10-12 weeks of age, increase in medial mass occurred with increase in the number of the smooth muscle cell layers whereas at 28 weeks further increase in media could be due to hypertrophy of the smooth muscle cells. In the small arteriolar vessels, medial enlargement was due at all ages to an increase in the number of smooth muscle layers. Our results show that in the SHR hypertrophy of the media occurs not only in the small arteriolar vessels, but also in large elastic and muscular arteries.

Animals↗
PubMed1983

Ultrastructural changes in mesenteric arteries from spontaneously hypertensive rats. A morphometric study.

Morphometric measurements at the electron microscope level were carried out on three categories of mesenteric arteries representing elastic (superior mesenteric), muscular and arteriolar vessels, from 10- to 12-week-old spontaneously hypertensive rats (SHR) and age-matched Wistar-Kyoto normotensive rats (WKY). Changes were observed only in muscular and arteriolar vessels of SHR, mainly as thickening of the vessel wall due to hypertrophy of the media. In muscular arteries, hypertrophy of the endothelial cells, widening of the subendothelial space, increased volume of the internal elastic lamina (IEL), and both hyperplasia and hypertrophy of the smooth muscle cells (SMC) in the media contributed to the wall thickening. In arteriolar vessels, increase in the subendothelial space and IEL, and hyperplasia of the SMC in the media were involved in the increased thickness of the vessel wall. There was no difference in the collagen content in all vessels, but elastin was increased in the muscular and arteriolar vessels of SHR. Nerve density was also increased in arteriolar vessels of SHR. These changes, especially the increase of SMC in muscular and arteriolar vessels, may be related to the elevated blood pressure in SHR.

Animals↗
PubMed1983

Comparison of blood vessel wall dimensions in normotensive hypertensive rats by histometric and morphometric methods.

The histometric method is often used in hypertension studies in order to reconstruct, from cross-sectional profiles of vessels in the contracted state, a hypothetical relaxed state of the vessels based on the measured length of the internal elastic lamina (IEL). In this study, the accuracy of the histometric method was tested by comparing values obtained with the histometric method from contracted vessels versus values from morphometric measurements of vessels of the same calibre but maximally relaxed. Mesenteric arteries from spontaneously hypertensive and normotensive Wistar-Kyoto rats were used. We found that the lumen size of the vessels was dependent on the methods of measurement and preparation. Lumen size, determined with the histometric method from contracted vessels, tended to be underestimated mainly because the IEL became shortened during contraction of the vessels. When vessels were prepared by perfusion fixation, a high perfusion flow rate (7.6 ml/min) caused expansion of the lumen in muscular (lumen diameter 120--250 microns) and arteriolar vessels (40--100 microns), but not in the elastic arteries (400--700 microns). The cross-sectional area of the vessel wall, however, remained unchanged when the vessels were either contracted with norepinephrine or expanded with high transmural pressure, so morphometric measurements of the vessel wall and its components can be done on both contracted and relaxed vessels. Our results suggest that the use of the histometric method for the estimation of relaxed lumen size from the contracted vessels is of very limited value. We also suggest that, for an accurate comparison of vessel wall dimensions between hypertensive and normotensive animals, cross-sectional area should be used instead of wall thickness alone.

Animals↗
PubMed1983

Biochemical and structural characterization of membrane fractions from bovine lens.

Plasma membranes of vertebrate lens fiber cells contain large numbers of gap junctions that may provide pathways for metabolic cooperation. Characterization of fiber cell gap junctions is thus necessary to understand this function. In this study, plasma membrane fractions were isolated from bovine lens according to established techniques, but without urea, detergents, or proteolytic enzymes. Electron microscopy indicated that isolated plasma membranes with gap junctions form double-membrane vesicles, and gap junctions comprised approximately 35% of the total membrane area in the crude fraction. These vesicles were impermeable to cationized ferritin, suggesting that they were sealed, and may be useful for permeability studies. Treatment of the crude fraction with 2.5% beta-mercaptoethanol or dithiothreitol caused reversible separation of junctional membranes, suggesting that disulfide bonds may be important in maintaining gap junction structure. Fractions with varying proportions of gap junctions were isolated using linear sucrose density gradient centrifugation. The proportional area of gap junction membrane versus total membrane in the various fractions ranged from 10% to at least 51%. The following plasma membrane enzymes were assayed in all fractions: Mg++-ATPase, Ca++-ATPase, alkaline phosphatase, phosphodiesterase, 5'-nucleotidase, and Na+, K+-ATPase. There was no correlation between enzyme activity and gap junction enrichment. This suggests that these enzymes are not associated with fiber cell gap junctions.

5'-Nucleotidase↗
PubMed1983

Effects of 4-aminopyridine and tetraethylammonium chloride on the electrical activity and cable properties of canine tracheal smooth muscle.

The actions of 4-aminopyridine (4-AP) on passive membrane properties of canine tracheal smooth muscle were compared to those of tetraethylammonium chloride (TEA). Transmembrane potential was studied with intracellular electrodes in a partitioned bath by extracellular current injection. The cells were electrically quiescent and showed rectification to depolarizing pulses. TEA (30 mM) depolarized the muscle and increased membrane resistance, time constant (282-421 msec), space constant (2.16-2.67 mm), decreased rectification and generated action potentials. D600, with TEA, abolished the spikes but further increased the membrane resistance, time constant (690 msec) and space constant (3.79 mm). TEA apparently blocked potassium conductance and initiated Ca++-dependent spikes owing to depolarization. 4-AP (5 mM) depolarized the membrane, but caused slow oscillations, decreased membrane resistance and space constant (1.94 mm), without affecting the time constant (265 msec) or rectification. These membrane effects of 4-AP were reversed to a large extent by atropine (10(-7) to 10(-6) M). D600 blocked the 4-AP-induced oscillations but not the decreased membrane resistance or space constant. 4-AP produced characteristic electrical and mechanical responses in a sucrose gap chamber when the muscle was functionally denervated by tetrodotoxin or scorpion venom. 4-AP appeared to activate the muscarinic receptor of canine trachealis to increase conductance, possibly to Na+ and K+. An effect on potassium conductance blockade might have been masked by the muscarinic receptor gated conductance changes, but is unlikely as it was not evident after exposure to atropine. We conclude that 4-AP actions in canine trachealis reflect chiefly activation of muscarinic receptors and not interference with potassium conductance.

4-Aminopyridine↗
PubMed1982

Endocrine, structural, and functional changes in the uterus during premature labor.

Rats which were subjected to ovariectomy on day 18 of pregnancy and were treated with 17 beta-estradiol underwent delivery prematurely at 0900 +/- 1.9 hours on the morning of day 18. All animals had in plasma and uterine tissue a precipitate and highly significant progesterone withdrawal and a corresponding significant increase in prostaglandin F and prostaglandin F metabolite. Progesterone replacement therapy given to a second group of castrated animals prevented progesterone withdrawal and premature labor, because the hormonal profile in plasma and uterine tissue of these animals was identical with that of the intact pregnant vehicle controls. Electron microscopy of longitudinal and circular myometrial layers showed a precipitate and highly significant increase in the number, size, and area of gap junctions in the uteri of the group undergoing premature delivery. In the uteri of the progesterone-treated and vehicle control groups (both intact pregnant), gap junctions were conspicuously scarce. Thus the extensive regulatory imbalance, provoked by progesterone withdrawal, induced a significant increase in myometrial gap junctions. This structural change established contacts between individual myometrial cell which could transform the multibillion cell community of the uterus into a syncytium, to generate low-resistance pathways to the flow of current and thus promote the propagation of trains of electrical discharge in support of labor.

Animals↗
PubMed1982

Improved electrical coupling in uterine smooth muscle is associated with increased numbers of gap junctions at parturition.

We have studied some passive electrical properties of uterine smooth muscle to determine whether a change in electrical parameters accompanies gap junction formation at delivery. The length constant of the longitudinal myometrium increased from 2.6 +/- 0.8 mm (X +/- SD) before term to 3.7 +/- 1 mm in tissues from delivering animals. The basis of the change was a 33% decrease in internal resistance and a 46% increase in membrane resistance. Axial current flow in an electrical syncytium such as myometrium is impeded by the cytoplasm of individual cells plus the junctions between cells. Measurement of the longitudinal impedance indicated that the specific resistance of the myoplasmic component was constant at 319 +/- 113 omega . cm before term and 340 +/- 93 omega . cm at delivery. However, a decrease in junctional resistance was apparent from 323 +/- 161 omega . cm to 134 +/- 64 omega . cm at delivery. 1.5-2 d after delivery, the junctional resistance was increased, as was the myoplasmic resistance. Thin-section electron microscopy of some of the same muscle samples showed that gap junctions were present in significantly greater numbers in the delivering tissues. Therefore, our results support the hypothesis that gap junction formation at delivery is associated with improved electrical coupling of uterine smooth muscle.

Action Potentials↗
PubMed1982

Changes in hormone levels and gap junctions in the rat uterus during pregnancy and parturition.

To study the temporal relationship between changes in hormone levels with the development of myometrial gap junctions, rats were sacrificed to collect uterine vein blood and uterine tissues daily from Day 15 to Day 20 of pregnancy, every 3 h on Day 21, during delivery, and 3 h after parturition. Levels of progesterone, estradiol and prostaglandin F2 alpha (PGF2 alpha) were measured by specific radioimmunoassay methods, and myometrial gap junctions were measured by quantitative electron microscopy. Levels of progesterone declined markedly on Day 19 of pregnancy, at least 60 to 70 h before the expected time of parturition and the decrease was more rapid in uterine tissue than in plasma. Levels of estradiol started to increase after Day 19, showed a marked rise on Day 21, and peaked at the time of parturition. Levels of PGF2 alpha also increased rapidly after Day 20, when progesterone levels had already dropped by 67% and 84% of Day 15 levels in plasma and tissue samples, respectively. Gap junctions, identified as 5- or 7-lined structures with a gap of about 2 nm at 100,000X magnification were present in the pregnant uterus. However, their number, size and area were increased significantly on Day 21 and during parturition. Increased numbers of gap junctions were still present in rats killed 3 h after parturition. These studies demonstrate that a decrease in progesterone levels followed by increases in estradiol and PGF2 alpha are coincident with the formation of gap junctions which in turn may coordinate the increased uterine activity required for parturition.

Animals↗
PubMed1982

Effects of glutaraldehyde fixative osmolarities on smooth muscle cell volume, and osmotic reactivity of the cells after fixation.

The contribution of glutaraldehyde (GA) to the effective osmolarity of GA fixatives, the osmotic reactivity of the cells after fixation in GA, and also the duration of fixation in GA on cell volume, were investigated using cultured smooth muscle cells (SMC) and spiral aortic strips. Four fixation procedures were studied. We found that GA contributes to the total effective osmolarity of the fixatives, and that the type of buffers used for the fixatives can also affect the cell volume differently during GA fixation. After GA fixation, the cells were still osmotically reactive, regardless of the buffer types for making up the GA fixatives, so that the osmolarity of the wash buffer after GA fixation is important. However, OsO4 eliminates osmotic responses, thus the osmolarity of OsO4 fixative and wash buffer have negligible influence on the cell volume. Longer fixation time up to 4 h had no effect of the cell volume.

Aldehydes↗
PubMed1982

Changes of gap junctions in myometrium of guinea pig at parturition and abortion.

Myometrial tissues from guinea pigs were quantitatively examined for gap junctions in electron micrographs. Small numbers of gap junctions were present between smooth muscle cells in myometria of pregnant guinea pigs at days 50 and 65 of gestation. The junctions increased in number and size at parturition on day 69 and decreased again to control levels 24 h after parturition. A similar increase in junctions occurred when abortion was induced by 16,16-dimethylprostaglandin E2 (PGE2) on day 65. There were no consistent or significant differences in number of gap junctions from myometrium taken over sites of placental attachment and from other sites. These results together with previous studies suggest that an increase in myometrial gap junction area is associated with and may be essential for parturition in guinea pigs, but the control of their development may differ from that in other mammals.

Abortion, Induced↗
PubMed1982

Ovarian hormones are not necessary for postpartum regression of gap junctions.

We tested the hypothesis that gap junction degradation in myometrium after parturition in rats requires an ovarian hormone. Compared with unoperated or sham-operated controls, ovariectomy on day 21 slightly delayed parturition (nearly all animals delivered on day 23 instead of day 22). It also significantly reduced the number and membrane area occupied by gap junctions in delivering animals. However, it did not alter the disappearance of gap junctions which occurs less than 24 h after parturition or disassociate gap junction disappearance from parturition. These results do not support the hypothesis of a necessary role for ovaries in gap junctions degradation after parturition and suggest that events associated with parturition may initiate this event.

Animals↗