Review Article
Mg-ATP binding: its modification by spermine, the relevance to cytosolic Mg2+ buffering, changes in the intracellular ionized Mg2+ concentration and the estimation of Mg2+ by 31P-NMR
- Daniel Lüthi, Dorothee Günzel, John A. S. McGuigan
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- 03 January 2001, pp. 231-252
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It is now generally accepted that the intracellular ionized magnesium concentration ([Mg2+]i) in muscle cells is around 1 mmol l-1; in heart muscle this means that from the total some 90-95 % is bound (see McGuigan et al. 1991a). Although binding will include sequestration by intracellular organelles, a large part of the binding is by ATP in the cytosol and an equilibrium exists in the cytosol between free ATP, ionized magnesium and Mg-ATP. The extend of this equilibrium depends on the equilibrium constant of the reaction, which is a function of pH, temperature and ionic strength. This equilibrium constant is also important in the estimation of [Mg2+]i using 31P-NMR. In this method the difference between the α and β peaks of ATP is measured and from this shift and the equilibrium constant between Mg2+ and ATP in the cytosol, the [Mg2+]i can be calculated (Vink, 1993).
Cardiac electrophysiological effects of platelet-derived substances
- N. A. Flores, A. N. S. Botchway, B. M. Stavrou, D. J. Sheridan
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- 03 January 2001, pp. 253-274
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The ability of the heart to function as an efficient pump is critically dependent on an adequate blood supply to the myocardium. When myocardial blood supply is reduced (ischaemia), contractile function of the heart is impaired and its cellular electrophysiological properties are profoundly altered. Prolonged ischaemia may result in cell necrosis which, if irreversible, leads to cell death (infarction) and predisposes the heart to ventricular arrhythmias, which in turn may be fatal. In man, myocardial ischaemia is the consequence of narrowing or occlusion of a coronary artery and is usually a manifestation of coronary artery disease which is often associated with thrombosis.
Research Article
H+-zwitterionic amino acid symport at the brush-border membrane of human intestinal epithelial (CACO-2) cells
- David T. Thwaites, Beverley C. Stevens
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- 03 January 2001, pp. 275-284
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Transport of a number of dipolar amino acids (and the orally active antibiotic D-cycloserine) across the apical membrane of human intestinal epithelial (Caco-2) cell monolayers is mediated by a Na+-independent, pH-dependent transport mechanism. Relatively little is known about the mode of action of this transport system so to differentiate between pH dependence and proton coupling three experimental protocols were designed and tested. The results demonstrate, firstly, that it is the transapical pH gradient and its maintenance (rather than apical acidity alone) that is important in amino acid uptake. Secondly, Na+-independent uptake of seven dipolar amino acids (with pKa (-log of acid dissociation constant) values between 1.50 and 4.23) showed a similar dependence on apical pH (half-maximal uptake being observed at pH 5.99-6.20). Thirdly, the pattern of pH-dependent amino acid (β-alanine) uptake is similar irrespective of whether the cationic substrate concentration is varied or constant, demonstrating no relationship between uptake and concentration of the cationic form of the amino acid. These observations demonstrate that the transport mechanism is a H+-zwitterionic amino acid symporter and suggest that the presence of a H+ gradient at the apical surface of the human small intestine (in the form of the acid microclimate) may be important in driving nutrient absorption.
Drug extrusion, 125I- efflux and the control of intracellular [Ca2+] in drug-resistant ovarian epithelial cells
- H. L. McAlroy, D. L. Bovell, J. A. Plumb, P. Thompson, S. M. Wilson
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- 03 January 2001, pp. 285-297
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Experiments were undertaken using an ovarian adenocarcinoma cell line (A2780) and a drug-resistant strain (A2780.ad) derived from this line. P-glycoprotein could not be detected in A2780 cells but was essentially ubiquitous in A2780.ad cells, although removing the selective pressure for drug resistance led to reduced expression. However, the amount of P-glycoprotein present was used to predict the capacity of these cells to extrude rhodamine-123 (R-123) and their resistance to adriamycin, a cytotoxic drug. This accords with the role of P-glycoprotein as a drug pump. Although hypotonic solutions increased anion efflux from A2780 and A2780.ad cells, larger responses occurred in the parental line. Moreover, R-123 extrusion and anion efflux appeared to be mutually independent processes and so these data do not support the view that P-glycoprotein is involved in the control of volume-sensitive anion channels. Hypotonic solutions increased intracellular free calcium ([Ca2+]i) in drug-resistant cells but not in the parental line, and so establishing a drug-resistant strain may affect the control of [Ca2+]i during osmotic swelling. This could account for effects that were previously attributed to P-glycoprotein.
Interaction of islet hormones with cholecystokinin octapeptide-evoked secretory responses in the isolated pancreas of normal and diabetic rats
- Jaipaul Singh*, Ernest Adeghate, Gines M. Salido, Jose A. Pariente, Maria D. Yago, Lubna O. M. Juma
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- 03 January 2001, pp. 299-318
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This study investigates the effects of the islet hormones, insulin (Ins), glucagon (Glu) and somatostatin (Som) with cholecystokinin octapeptide (CCK-8) on amylase secretion and intracellular free calcium concentration [Ca2+]i and their pattern of distribution in the isolated pancreas of normal and diabetic rats. Ins and Glu evoked small increases in amylase output from pancreatic segments compared with a much enhanced effect of CCK-8. In contrast, Som induced a biphasic response comprising an initial decrease followed by a secondary increase and this biphasic response may be dependent upon the concentration. Combining the islet hormones with CCK-8 resulted in marked potentiation in amylase output compared with either CCK-8 alone or the individual hormone. Genistein and tyrphostin A25, the tyrosine kinase inhibitors, evoked a small decrease in amylase output from pancreatic segments. They had no effect on the CCK-8-evoked secretory response but markedly inhibited the potentiation of the islet hormones with CCK-8. In pancreatic acini and acinar cells Ins, Glu and Som individually evoked small increases in amylase output compared with a much larger response with CCK-8. When the islet hormones were combined with CCK-8 there was no potentiation of amylase output. Similarly, when rats were rendered diabetic by prior treatment with streptozotocin Ins, Glu and Som failed to potentiate the secretory response of CCK-8. In fura-2-loaded pancreatic acinar cells Ins or Glu evoked small increases in [Ca2+]i compared with a much larger elevation with CCK-8. Ins, Glu and Som each enhanced the CCK-8-evoked [Ca2+]i. Genistein elicited a decrease in [Ca2+]i both in the absence and presence of the islet hormones. It also decreased the elevation in [Ca2+]i resulting from the combined presence of CCK-8 with either Ins or Glu but it had no effect on CCK-8 in combination with Som. In pancreatic acinar cells from diabetic rat Ins, Glu and Som had no detectable effect on CCK-8-evoked elevation in [Ca2+]i compared with the response obtained with CCK-8 alone. CCK-8-immunopositive cells were distributed around the walls of blood vessels, numerous Ins-positive cells in the central and peripheral parts of the islets of Langerhans, Glu-immunoreactive cells in the periphery of islets and Som-positive cells in the outer part of the islets. During diabetes, the number of CCK-immunopositive cells remained unchanged whereas the number of Ins-positive cells decreased coupled with an increase in the number of Glu-positive cells. The results indicate that both tyrosine kinase and cellular Ca2+ seem to be the intracellular mediators involved with the enhanced secretory responses obtained with a combination of the islet hormones with CCK-8. Moreover, the presence of viable pancreatic islets of Langerhans seems to be associated with the potentiation of the islet hormones with CCK-8.
Role of nitric oxide in indomethacin-induced gastric mucosal dysfunction in the rat
- Vildan Gürbüz, Inci Alican, Berrak Ç. Yegen, Ayhan Bozkurt, Berna Oktar, Goncagül Haklar, Meral Yüksel, Hizir Kurtel
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- 03 January 2001, pp. 319-332
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The present study was undertaken to explore the role of nitric oxide (NO) in the pathogenesis of experimental non-steroidal anti-inflammatory drug (NSAID)-induced gastropathy. We assessed the role of NO inhibition and donation in indomethacin-induced gastric mucosal dysfunction. The stomach was perfused with vehicle (control) for 20 min, followed by indomethacin (10 mg ml-1 in 1.25 % sodium bicarbonate, pH 8.4) for 120 min. NG-nitro-L-arginine methyl ester (L-NAME, 5 and 10 mg kg-1, I.V. bolus), L-arginine, D-arginine (100 mg kg-1I.V. bolus, 10 mg kg-1 h-1, 2 h infusion) and the NO donor glyceryl trinitrate (GTN) were given at the same time (20, 40 and 80 µg kg-1 min-1, 15 min infusion) as perfusion with indomethacin was started. Epithelial permeability was quantified by measuring blood-to-lumen clearance of 51Cr-labelled EDTA. Indomethacin caused a 20-fold increase in 51Cr-EDTA leakage compared with that of the control group. Treatment with L-NAME or L-arginine did not affect the indomethacin-induced alterations in mucosal permeability. Administration of GTN (20 µg kg-1 min-1) significantly reduced the indomethacin-induced mucosal dysfunction. By contrast, higher doses of GTN (80 µg kg-1 min-1) exacerbated epithelial dysfunction induced by indomethacin. Elevated levels of carbonyls and myeloperoxidase (MPO) observed after indomethacin administration were significantly reduced, to the control values, when GTN (20 µg kg-1 min-1) was administered along with indomethacin. These data suggest that NO from exogenous sources can exert a dual action on the integrity of the gastric mucosa challenged by indomethacin. Low doses of GTN can prevent mucosal dysfunction induced by indomethacin, while higher doses of GTN may exacerbate the increases in epithelial permeability.
Regulatory role of cAMP in transport of Na+, Cl- and short-chain fatty acids across sheep ruminal epithelium
- Gotthold Gäbel*, Heidrun Butter, Holger Martens
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- 03 January 2001, pp. 333-345
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Sodium is absorbed in considerable amounts across the ruminal epithelium, whilst its transport is strongly interrelated with the permeation of chloride and short-chain fatty acids (SCFAs). However, regulation of ruminal Na+, Cl-, and SCFA absorption is hardly understood. The present study was therefore performed to characterize the influence of cAMP on sodium and sodium-coupled transport mechanisms in short-circuited, stripped ruminal epithelia of sheep. Elevation of intracellular cAMP concentrations by theophylline (10 mM) or theophylline in combination with forskolin (0·1 mM) significantly reduced mucosal-to-serosal sodium transport, leading to a reduction of net transport. The theophylline- or theophylline-forskolin-induced reduction of sodium transport was accompanied by a decrease in chloride net transport but revealed no effect on propionate flux. Short-chain fatty acids stimulated Na+ transport but their stimulatory effect was almost completely blocked by theophylline-forskolin. In solutions with and without SCFAs, the inhibitory effect of 1 mM amiloride on sodium transport was strongly reduced after theophylline-forskolin pretreatment of the tissues. Blocking the production of endogenous prostaglandins by addition of indomethacin (10 µM) led to a theophylline-sensitive stimulation of unidirectional and net fluxes of sodium. The findings indicate that apical, amiloride-sensitive Na+-H+ exchange and/or basolateral Na+-K+-ATPase can effectively be blocked by cAMP, leading to a decrease in sodium and chloride transport. In the ruminal epithelium, cAMP is a second messenger of prostaglandins, which are released spontaneously under in vitro conditions.
Mechanically induced potentials in fibroblasts from human right atrium
- A. Kamkin, I. Kiseleva, K. D. Wagner*, A. Lammerich, J. Bohm, P. B. Persson, J. Günther
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- 03 January 2001, pp. 347-356
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It has been shown that cardiac fibroblasts of the human heart are electrically non-excitable and mechanosensitive. The resting membrane potential of these cells is -15.9 ± 2.1 mV and the membrane resistance is 4.1 ± 0.1 GΩ. Rhythmic contractions of the myocardium associated with stretch of the surrounding tissue produce reversible changes in the membrane potential of cardiac fibroblasts. These mechanically induced potentials (MIPs) follow the rhythm of myocardial contractions. Simultaneous recording of the action potential of cardiomyocytes and MIPs of cardiac fibroblasts demonstrates a delay of 40.0 ± 0.4 ms after the action potential before the appearance of the MIP. Contraction produces a MIP which is more positive or more negative than the reversal potential - the membrane potential due to current injection at which the MIP reverses its direction. Regardless of the initial orientation of the MIP, intracellular polarization increases the amplitude towards the reversal potential if the background MIP had depolarized the membrane or away from the reversal potential if the initial background MIP had hyperpolarized the membrane. Artificial intracellular polarization changed the amplitude but not the frequency of the MIP. The pool of electrically non-excitable mechanosensitive cells, which change their electrical activity during contraction and relaxation of the heart, may play a role in the mechano-electrical feedback mechanism which has to be taken into account in the normal function of the heart as well as in pathological processes.
Effects of acute and chronic starvation on central and peripheral noradrenaline turnover, blood pressure and heart rate in the rat
- S. El Fazaa, L. Somody, N. Gharbi, A. Kamoun, C. Gharib, G. Gauquelin-koch
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- 03 January 2001, pp. 357-368
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When faced with stress, an organism calls upon several mechanisms to maintain biological homeostasis. The cardiovascular system is the first to respond usually with an increase in arterial pressure and tachycardia. Therefore we investigated the central and peripheral sympathetic responses to acute and chronic starvation in Wistar rats. The noradrenaline (NA) turnover rate was determined in different catecholaminergic nuclei (A1, A2, A5, A6) as well as the arterial blood pressure and heart rate modifications. During acute starvation (3 days of starvation), the NA turnover was increased in the A1 and rostral A2 nuclei as well as in ventricles and kidneys and decreased in the A6 nucleus. During chronic starvation (4 consecutive cycles of 3 days of starvation plus 1 day of feeding), the NA turnover was increased in the A5 and caudal A2 nuclei as well as in ventricles and atria and decreased in the A1 nucleus and kidneys. The arterial blood pressure revealed a gradual decrease during the first 3 days of fasting but the heart rate was not modified. We conclude that starvation should be considered as an unusual state of stress because of the absence of locus coeruleus response (A6 nucleus) despite its well-defined role in stress reactions. One of the manifestations of these central and peripheral noradrenergic changes is the change in blood pressure during the starvation-feeding cycles.
Effects of skin surface cooling and heating on autonomic nervous activity and baroreflex sensitivity in humans
- Hideki Kinugasa, Kaname Hirayanagi
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- 03 January 2001, pp. 369-377
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The influence of skin surface cooling and heating on heart rate variability (HRV), blood pressure variability (BPV), and baroreflex sensitivity (BRS) were studied in 11 healthy supine volunteers in air temperatures of 18°C (cool), 24°C (mild), 48°C (warm), and 60°C (hot) in a styrene foam chamber. The high-frequency component of HRV (HFRR) decreased and the ratio of low- to high-frequency components, LFRR/HFRR, increased with skin surface heating. Therefore, a suppression of cardiac vagal nervous activity and an enhancement of cardiac sympathetic nervous activity can be caused by skin surface heating. The low-frequency component of BPV (LFSBP), i.e. the fluctuations of the Mayer waves, increased with skin surface heating. The responses between the very low-frequency components of HRV (VLFRR) and systolic BPV (VLFSBP) to thermal skin stimulation were different, although these variables both were considered to be indicators of the thermoregulatory vasomotor control or renin-angiotensin system. Baroreflex sensitivity (BRS) did not increase with skin surface heating, at least in humans.
Isometric and concentric performance of electrically stimulated ankle plantar flexor muscles in intact rat
- Mark E. T. Willems, William T. Stauber
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- 03 January 2001, pp. 379-389
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The relationship between muscle force and ankle position during isometric and pre-loaded slow concentric contractions (angular velocity, 0.52 rad s-1; range of motion, 1.22 rad) and the recovery of isometric force following concentric contractions at different velocities were determined for electrically stimulated plantar flexor muscles in intact rats. Pre-loaded refers to the isometric contraction which immediately precedes the concentric contraction. Ankle position was controlled by a dynamometer and force was recorded under the sole of the foot. The peak isometric force (19.2 N) was nearly constant at all ankle positions (range of motion, 1.57 rad). The muscle length and distal fibre length of gastrocnemius medialis at ankle positions between 0.79 rad and 2.01 rad were increased by 12.6 % and 20.3 %, respectively. During slow concentric contractions, the force progressively decreased (23.1 ± 2.1 %); the force decreased by only 6.3 ± 0.9 % during sustained isometric contractions of similar duration (3400 ms). The recovery of isometric force following concentric contractions with similar stimulation frequencies (80 Hz) was velocity dependent (i.e. more rapid at higher velocities). It is concluded that pre-loaded slow concentric contractions of the plantar flexor muscles in intact rats do not follow the same relationship as that of isometric force and ankle position. Our results in intact rats show that the force output of electrically stimulated ankle plantar flexor muscles measured under the sole of the foot can be used to study the physiological properties of skeletal muscle working in situ.
The use of a deuterium tracer technique to follow the fate of fluids ingested by human subjects: effects of drink volume and tracer concentration and content
- C. P. Lambert, D. Ball, J. B. Leiper, R. J. Maughan
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- 03 January 2001, pp. 391-399
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Deuterium oxide (2H2O) has been added to drinks as a tracer for water to estimate the availability to the body water pool of ingested fluids, but doubts have been raised as to the reliability of the method. The present investigation evaluated the effects of systematic variations in the volume of fluid consumed and the amount and concentration of added tracer on the rate of accumulation of tracer in arterialized blood after ingestion of a labelled drink. Three separate experiments were undertaken. In expt 1, six healthy men ingested on separate occasions 200, 400 and 800 ml of a dilute glucose-electrolyte solution: all test drinks contained the same concentration (40 g l-1) of 2H2O. In expt 2, six healthy men ingested 200, 400 and 800 ml of the same glucose-electrolyte drink: each drink contained 8 g of 2H2O so that the concentration, but not the amount, of 2H2O differed between treatments. In expt 3, six healthy men ingested 400 ml of the same drink on three separate occasions: each drink contained 8, 16 or 32 g of tracer so that amount and concentration of 2H2O both varied. Arterialized venous blood samples were collected for the determination of deuterium (2H) concentration before ingestion of the test drink and at intervals for 120 min after ingestion. All trials for each of the experiments were conducted in the morning after an overnight fast and trials were in randomized order and separated by 7 days. In expt 1, the blood 2H concentration at all time points from 2 min after ingestion of the test drink onwards was higher for the drink containing 32 g 2H2O than for the drink containing 16 g 2H2O, which in turn was higher than after ingestion of the drink containing 8 g of 2H2O. In expt 2, no significant differences between treatments were observed at any time. In expt 3, the rate of 2H accumulation was greater after ingestion of the drink containing 32 g of 2H2O than after either of the other two drinks, and the 2H accumulation rate was greater after ingestion of the drink containing 16 g of 2H2O than after the drink containing 8 g of 2H2O. When data from all three experiments were combined, significant correlations were observed between the rate of accumulation of 2H in the circulation (p.p.m. min-1) and the amount (rs = 0.75, P < 0.001) and concentration (rs = 0.69, P < 0.001) of 2H2O in the test drink, but there was no relationship (rs = 0.09, P = 0.5) between the rate of 2H accumulation in the blood and the volume of the drink consumed. The results suggest that the rate of tracer accumulation in the blood after ingestion of different volumes of test drinks is not a reliable indication of the availability of the ingested fluid, but that the method gives at least a qualitative measure of the sum of the effects of gastric emptying and intestinal water absorption.
Sweat ethanol concentrations are highly correlated with co-existing blood values in humans
- Michael J. Buono
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- 03 January 2001, pp. 401-404
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This study compared the concentration of ethanol, both absolute and relative to water content, in sweat and blood. Ten male volunteers consumed approximately 13 mmol (kg body weight)-1 of ethanol. Blood and sweat samples were collected approximately 1, 2 and 3 h following ingestion. Sweat was collected following pilocarpine iontophoresis using an anaerobic technique that prevented ethanol evaporation. In addition, the water content of sweat and blood samples was determined. The correlation between sweat and blood ethanol, expressed in mmol l-1, was r = 0.98. The slope of the relationship was 0.81. When corrected for the water content in each sample, and expressed as mmoles per litre of water, the correlation remained very high (r = 0.97) while the slope increased to 1.01. These results suggest that rapid and complete equilibrium of ethanol occurs across the sweat gland epithelium.
Comparison of microvascular filtration in human arms with and without postmastectomy oedema
- A. W. B. Stanton*, B. Holroyd, P. S. Mortimer, J. R. Levick
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- 03 January 2001, pp. 405-419
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Oedema is caused by impaired lymphatic drainage and/or increased microvascular filtration. To assess a postulated role for the latter in postmastectomy oedema, filtration was studied in the forearms of 14 healthy subjects and 22 patients with chronic, unilateral arm oedema caused by surgical and radiological treatment for breast cancer. A new non-contact optical device (the Perometer) and a conventional mercury strain gauge were used simultaneously to record forearm swelling rates caused by microvascular filtration during applied venous congestion. Filtration rate (FR) per 100 ml tissue was measured over 10-15 min at a venous pressure of 30 cmH2O, a pressure reached in the dependent forearm (FR30), and then at 60 cmH2O (FR60). Apparent filtration capacity of 100 ml soft tissue (CFCa) was calculated from FR60 - FR30/30, after adjustment for bone volume. The Perometer and strain gauge gave similar results in normal and oedematous arms. Mean CFCa in healthy subjects was (3·8 ± 0·4) × 10-3 ml (100 ml)-1 cmH2O-1 min-1, close to literature values. In the patients, FR30 was 47 % lower in the oedematous forearm than in the opposite, unaffected forearm (P = 0·04). FR60 showed a similar trend but did not reach significance (P = 0·15). The values of CFCa of (2.2 ± 0.5) × 10-3 ml (100 ml)-1 cmH2O-1 min-1 in the oedematous arm and (2.8 ± 0.5) × 10-3 ml (100 ml)-1 cmH2O-1 min-1 in the unaffected arm were not significantly different (P = 0.47). When differences in arm volume on the two sides were taken into account, the total fluid load on the lymphatic system of the oedematous forearm was (411.0 ± 82.2) × 10-3 ml min-1 at 30 cmH2O and (1168 ± 235.6) × 10-3 ml min-1 at 60 cmH2O, similar to the normal side, namely (503.7 ± 109.3) × 10-3 ml min-1 and (1063 ± 152.0) × 10-3 ml min-1, respectively (P >= 0·50). The filtration capacity of the entire oedematous forearm (CFCa scaled up by total soft tissue volume), (25.4 ± 6.2) × 10-3 ml cmH2O-1 min-1, was not significantly greater than that of the normal forearm, (18.3 ± 2.6) × 10-3 ml cmH2O-1 min-1 (P = 0.40). The results indicate that no major change occurs in the microvascular hydraulic permeability-area product of the forearm, or in the total filtration load on the lymph drainage system during dependency, in the arm with postmastectomy oedema compared with the normal arm. This argues against a significant haemodynamic contribution to postmastectomy oedema.
Breast growth and the urinary excretion of lactose during human pregnancy and early lactation: endocrine relationships
- David B. Cox, Jacqueline C. Kent, Tammy M. Casey, Robyn A. Owens, Peter E. Hartmann
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- 03 January 2001, pp. 421-434
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Breast volume and morphology of eight subjects were measured before conception and at intervals throughout pregnancy until 1 month of lactation. Breast volume before conception ranged from 293 to 964 ml. At the end of pregnancy the volume of breast tissue had increased by 145 ± 19 ml (mean ± S.E.M., n = 13 breasts, range 12-227 ml) with a further increase to 211 ± 16 ml (n = 12 breasts, range 129-320 ml) by 1 month of lactation. Urinary excretion of lactose increased at 22 weeks of pregnancy, signalling the capacity of the breast to synthesize lactose at this time. During pregnancy, both the change in breast volume and the change in cross-sectional area of the areola were related to the concentration of human placental lactogen in the plasma. The growth of the nipple and the rate of excretion of lactose were related to the concentration of prolactin in the plasma. During the first 3 days after birth, the rate of excretion of lactose was related to the rate of excretion of progesterone. There was no relationship between the growth of the breast during pregnancy and the amount of milk produced at 1 month of lactation.
Breast volume and milk production during extended lactation in women
- Jacqueline C. Kent, Leon Mitoulas, David B. Cox, Robyn A. Owens, Peter E. Hartmann
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- 03 January 2001, pp. 435-447
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Quantitative measurements were made of relative breast volume and milk production from 1 month of lactation until 3 months after weaning, and the storage capacity of the breasts was calculated. The increase in breast tissue volume from before conception until 1 month of lactation was maintained for the first 6 months of lactation (means ± S.E.M.) (190.3 ± 13.1 ml, number of breasts, nb = 46). During this period of exclusive breast-feeding, 24 h milk production from each breast remained relatively constant (453.6 ± 20.1 g, nb = 48), and storage capacity was 209.9 ± 11.0 ml (nb = 46). After 6 months, breast volume, milk production and storage capacity all decreased. There was a relationship between 24 h milk production and the storage capacity of the breasts, and these both appeared to be responding to infant demand for milk. At 15 months of lactation, the 24 h milk production of each breast was substantial (208.0 ± 56.7 g, nb = 6), even though the breasts had returned to preconception size. This was associated with an apparent increased efficiency of the breast (milk production per unit breast tissue) after 6 months, which may have been due to redistribution of tissues within the breast. The possible causes of the decrease in breast volume are discussed.