Structure and mechanics of growing arterial microvessels from hypertrophied urinary bladder in the rat

Rat bladder hypertrophy, induced by a partial ligation of the urethra, was used to study the accompanying changes of microvascular smooth muscle mechanics, pharmacology and morphology. A segment of a microarterial vessel to the bladder was taken from a defined anatomical location and studied in a wire myograph in vitro at the length for maximal isometric force development (L _max). After 10 days of ligation, bladder hypertrophy resulted in a microvascular growth response compared to non-operated controls which was characterized by (i) an increase of the calculated diameter at L _max from 134±5 m to 222±19 m; (ii) an increase of the media thickness from 22.4±1.9 m to 32.2±3.0 m; (iii) an increase of the active tension from 1.42±0.28 mN/mm to 3.06±0.33 mN/mm; (iv) no change of the wall/lumen ratio (from 0.83±0.10 to 0.79±0.15). Normalized length/force relations (active, passive and total) did not differ significantly between microarteries from control and hypertrophic bladders. Microvascular smooth muscle growth was also associated with a decreased sensitivity to K⁺-induced depolarization and an increased sensitivity to ₁-adrenergic stimulation. No differences were noted regarding the Ca²⁺ sensitivity of force during K⁺-induced depolarization. The results suggest that microvascular growth (1) is immediately and positively influenced by the organ growth; (2) results in a functional resetting of the microvascular segments towards larger diameters without gross morphological or mechanical alterations; and (3) is accompanied by pharmacological alterations of the smooth muscle reactivity.     

Re-acceleration of the down-regulated contraction kinetics in the rat tracheal smooth muscle

The contraction kinetics of smooth muscle show a down-regulation after the transient rise found during sustained contraction. We tried to find out therefore if the contraction kinetics of rat tracheal smooth muscle can be re-accelerated during sustained activation. A 2 s length vibration (100 Hz sinusoidal; amplitude=6% of the muscle length) produces an immediate fall in the force developed by the activated muscle. A biexponential function was fitted to the force recovery. The reciprocal of the time constant,t ₂, describing the slow component of force recovery, reflects the kinetics of contraction. The contraction kinetics reach their highest levels (t ₂=4.9±0.1 s,n=166) about 30 s after the onset of electrical field stimulation. Three experimental groups were activated by either 10 M serotonin (5-HT), 100 M acetylcholine (ACh), or by 2 M ACh for 50 min. Approximately 10 vibrations were applied to each preparation after an 8 min activation in order to observe stabilized down-regulated contraction kinetics.t ₂ values were calculated from the force recovery after vibration and averaged 11.2±0.2 s (n=141), 11.5±0.2 s (n=137), and 11.1±0.3 s (n=84), respectively. After 50 min of continuous chemical activation, the preparation was stimulated additionally by the neurogenic release of acetylcholine. Thet ₂ of post-vibration force recovery, as measured after 30 s of neural activation, showed no change in the specimens basically activated by 100 M ACh (11.0±0.4 s,n=51). A decline int ₂, indicating accelerated kinetics, was observed in the groups which had been stimulated by 10 M 5-HT (5.9±0.2 s,n=51) and 2 M ACh (5.6±0.2 s,n=47). The re-accelerating effect of the second stimulus could be reproduced recurrently. The down-regulated contraction kinetics can be re-accelerated either by activating another receptor type in addition to the one already maximally stimulated or by increasing the stimulus mediated by one of the receptor types from half maximal to maximal strength. However, this is only possible if the additional activation is strong enough, as indicated by an increase in active force. It could be demonstrated that the slowing of the cross-bridge cycling rate is the result of a regulatory process and not the result of substrate deficiencies or refractoriness in the regulatory of contractile proteins.     

Regulation of force in skinned,single cells of ferret aortic smooth muscle

An isolation technique was developed for single cells from the ferret aorta, which resulted in the isolation of long (87±27 m; x±SD, n=62), relaxed, pharmacologically active smooth muscle cells. These cells were attached to microtools, one of which was connected to a force transducer. Force in maximally phenylephrine-stimulated contractions of the intact cells averaged 2.3 ±1.4 N (n=17). After cell skinning with saponin, the threshold for force development was 0.05 M [Ca²⁺], and force reached a maximum of 4.4±1.6 N (n = 36) at 0.5 M [Ca²⁺]. Plots of relative steady-state force vs pCa (–log₁₀[Ca²⁺]) were fit to the Hill equation, which yielded a pCa at half-maximal force of 6.87 ± 0.30 and a Hill coefficient of 2.3±1.4 (n = 29). When 2.5 M calmodulin was added to the solutions, the calcium sensitivity of force was significantly increased (P<0.05) without changing the maximal force (P>0.05). In a solution of pCa 7, the skinned cells developed 2.5±0.5 N (n = 5) of force when stimulated with a phorbol ester. The addition of a specific inhibitor (17 kDa) of protein kinase C to the calcium buffers depressed (P<0.05) the maximally Ca²⁺ -activated force without a change in the calcium sensitivity of force (P>0.05). These data strongly suggest that in vascular smooth muscle, protein kinase C may be involved in a physiological, regulatory system for force.     

Mechanism of hyperthyroidism-induced modulation of the L-type Ca2+ current in guinea pig ventricular myocytes

The positive inotropic effects of thyroid hormone in the heart, increased force and velocity of contraction have been mostly attributed to modulation of myosin ATPase isoenzymes (V₁, V₂ and V₃), and sarcoplasmic reticulum Ca²⁺ pumping activity. In addition, we have suggested that the effects on ventricular contraction result from a thyroid hormone-induced increase in L-type Ca²⁺ current (I _{Ca, L}). Due to the central role of I _{Ca, L} in excitation-contraction coupling, we studied mechanisms whereby thyroid hormone augments this current. Since thyroid hormone modulates adenylate cyclase activity in various tissues, we tested the hypothesis that the hormone activates adenylate cyclase, leading to increased cyclic adenosine monophosphate (cAMP) levels, protein kinase A activation, Ca²⁺ channel phosphorylation and increased I _{Ca, L}. We therefore stimulated or inhibited different sites along the adenylate cyclase cascade, and measured I _{Ca, L} and isometric twitch in ventricular myocytes and papillary muscles from euthyroid and hyperthyroid guinea pigs. Our major findings were as follows. In euthyroid myocytes, 0.1 M isoproterenol (Iso) increased I _{Ca, L} (at V _M=0 mV) from –7.04±0.72 to –22.26±1.88 pA/pF, P<0.05, while in hyperthyroid myocytes (I _{Ca, L}=-21.48±2.94 pA/pF), Iso was ineffective. In euthyroid myocytes, intracellular application of cAMP (50 M) was as potent as Iso, but ineffective in hyperthyroid myocytes. In hyperthyroid myocytes, a protein kinase A inhibitor (2 M) lowered I _{Ca, L} from –26.82±1.54 to -10.17±1.70 pApF (P<0.05), but had no effect in euthyroid myocytes. In hyperthyroid myocytes, acetylcholine (ACh) (1 M) decreased I _{Ca, L} from –26.86±1.49 to –18.33±1.25 pA/pF (P<0.05), while in euthyroid myocytes ACh decreased I_{Ca, L} from –6.80±0.61 to –6.00±0.39 pA/pF (NS). Accordingly, in hyperthyroid papillary muscles, ACh decreased twitch tension by 36.4±2.8%, but in euthyroid preparations only by 9.4±5.1% (P<0.05). These findings suggest that thyroid-hormone-induced increase in I _{Ca, L} contributing to positive inotropy, is mediated by activation of the adenylate cyclase cascade.     

Modulation of bradykinin responses in airway smooth muscle by epithelial enzymes

We have studied the effect of epithelium removal on responses of guinea pig trachea to bradykinin (BK). BK (1 nM–10 M) gave a concentration-dependent relaxation when epithelium was present (E+: EC₅₀=10±3 nM). Epithelium removal resulted in a biphasic response to BK with relaxation at low concentrations (E–: EC₅₀=3.0±1.0 nM) and a recontraction to baseline at higher concentrations (EC₅₀=2.0±1 M). Phosphoramidon (10 M), an inhibitor of neutral endopeptidase (NEP), which cleaves BK into inactive peptides, potentiated relaxation (EC₅₀=1.0±0.9 nM and 0.1±0.1 nM in E+ and E respectively) and contraction in trachea with intact epithelium (EC₅₀=0.08±0.03 M). Inhibition of cyclooxygenase by indomethacin (5 M), inhibited relaxation to BK in E+ tracheal segments, resulting in a slight contraction (EC₅₀=1.0 M), whereas a potent contractile response was observed in E–segments (EC₅₀ 1.6 M, maximal contraction >1 g). In the presence of both indomethacin and phosphoramidon BK caused contraction, even in the presence of epithelium (EC₅₀=0.2±0.11 M), and the response in the absence of epithelium was similar to the response observed in trachea with intact epithelium (EC₅₀=0.25±0.1 M). The contractile effect of BK on airway smooth muscle may be inhibited by a protective role of epithelium, due to release of relaxant prostanoids and by degradation by epithelial NEP. In asthma, bronchoconstrictor responses to BK may be partly explained by loss of airway epithelium.     

Tubular transport processes in proximal tubules of hypothyroid rats. Lack of relationship between thyroidal dependent rise of isotonic fluid reabsorption and Na+−K+-ATPase activity

Hypothyroid rats reconstituted with 10 g/kg b.w. per day of tri-iodothironine (T₃) for 4 days resulted in normal free T₃ and TSH levels. FT₃ levels were: 0.53±0.3 pg/ml in hypothyroid rats; 2.78±1.21 pg/ml in hormone reconstituted rats and 2.90±0.90 pg/ml in euthyroid rats. TSH levels were 3,508±513 g/ml in hypothyroid rats; 1,008±204 g/ml in reconstituted rats and 270±184 ng/ml in euthyroid rats.When hypothyroid rats were reconstituted with 50 g T₃/kg b.w. per day, TSH levels were nearly normal after 4 days (1,157±621 ng/ml). However FT₃ levels after 1–4 days were always higher than in euthyroid rats.Hypothyroid rats show a decrease in isotonic fluid reabsorption (J _v) in the proximal tubule (1.50±0.08 versus 4.96±0.23 10^–2 nl·mm^–1·s^–1 in euthyroid animals). 1 day after T₃ (10 g/kg b.w./day) injectionJ _v was increased significantly to 2.05±0.20 10^–2 nl·mm^–1·s^–1 and continued to increase during 4 days of T₃ reconstitution.When 50 g T₃/kg b.w./day was used,J _v increased to 2.75±0.07 after 1 day and to 3.10±0.42 10^–2 nl·mm^–1·s^–1 after 4 days.J _v was never reaching a value close to that of euthyroid rats because the tubular radius in hypothyroid rats (14.7±1.8 m) is less than that of euthyroid rats (19.2±0.5 m). The radius in hypothyroid rats treated with T₃ was unchanged over a 4 day course with either high or low doses of T₃.Na⁺–K⁺-ATPase activity was found to be 2.91±0.16 M P_i/h×mg protein in homogenates of kidney cortex from hypothyroid rats. Treatment of hypothyroid rats with 10 g or 50 g of T₃ resulted in an initial decrease in ATPase activity, followed by an increase to base level in hypothyroid rats with 10 g and a significantly higher level with 50 g. This decrease in ATPase activity was contrasted to the increase inJ _v.These data indicate that there is a dissociation between the effects of physiological doses of thyroid hormones on proximal tubular reabsorption and the effects of T₃ on Na⁺–K⁺-ATPase activity of kidney cortex. This leads to question the relationship between sodium transport and ATPase activity under physiological doses of thyroid hormones. An early effect of physiological doses of thyroid hormones on brush border Na⁺ permeability is suggested.     

Muscle Proteolysis and Weight Loss in a Neonatal Rat Model of Sepsis Syndrome

Our hypothesis is that nitrogen loss in septic neonates is caused by increased muscle proteolysis. Sprague–Dawley rat pups (P7) were injected intraperitoneally with NaCl or 4 mg/kg/BW lipopolysaccharide (LPS) and then sacrificed at 2, 4, 24, and 48 hr. Sepsis syndrome was confirmed by elevated serum tumor necrosis factor (24.6 ng/mL ± 18.4 [LPS] and <1.0 ng/mL [controls]; p < .05). Proteolysis in gastrocnemius/soleus muscle was analyzed by quantitation of tissue tyrosine loss. The neonatal rats injected with LPS had significant media tyrosine release at 24 hr compared to the controls (0.39 ± 0.14 versus 0.25 ± 0.11 mol tyrosine/g muscle; p < .05). At 48 hr, LPS-induced muscle tyrosine release ceased (0.24 ± 0.04 [control] versus 0.23 ± 0.03 mol tyrosine/g muscle [LPS]). After 48 hr, gastrocnemium/soleus weight was less in the LPS-injected rats (50.5 ± 4.8 to 31.2 ± 4.0 g; p < .0001). Similar changes were not seen in the extensor digitorum longus, suggesting that some muscles were relatively preserved. Also, LPS resulted in significant weight loss. We conclude that selective muscle proteolysis contributes to nitrogen loss in neonatal sepsis. Although proteolysis abates by 48 hr, short-term injury results in significant muscle-mass deficit.     

Effects of diadenosine polyphosphates,ATP and angiotensin II on cytosolic Ca2+ activity and contraction of rat mesangial cells

Diadenosine polyphosphates (Ap _n A) are known to influence cellular Ca²⁺ activity ([Ca²⁺]_i) in several cells. Their vasoactive potency has been described in various systems including the kidney. We examined the effects of diadenosine polyphosphates, adenosine 5-triphosphate (ATP) and angiotensin II (Ang II) on cytosolic Ca²⁺ activity of mesangial cells (MC) in culture obtained from normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats. [Ca²⁺]_i was measured as a fluorescence ratio F ₃₄₀/F ₃₈₀ with the fura-2 technique using three excitation wavelengths (340 nm, 360 nm and 380 nm) and a photon counting tube. Resting [Ca²⁺]_i was not significantly different in MC from WKY and SHR rats and was measured as 132±9 nmol/l (n=65) and 114±12 nmol/l (n=36), respectively. Diadenosine polyphosphates (Ap₃A–Ap₆A) increased [Ca²⁺]_i transiently with an initial peak and a secondary plateau phase comparable to the effects of ATP or Ang II. Increases in [Ca²⁺]_i induced by all these agonists were not significantly different between MC of WKY and SHR rats. ATP, Ap₃A, Ap₄A, Ap₅A, Ap₆A (each 5 mol/l) increased the fura-2 fluorescence ratio initially by 0.66±0.09 (n=33), 0.52±0.08 (n=18), 0.25±0.05 (n=16), 0.09±0.06 (n=7), 0.09±0.04 (n=11), respectively. A half-maximal initial increase in the fura-2 fluorescence ratio was reached at 22 nmol/l, 0.9 mol/l, 2.0 mol/l and 4.0 mol/l with Ang II, Ap₃A, ATP and Ap₄A, respectively. Ap₄A (100 mol/l, n=18) led to a reversible contraction of MC. Diadenosine polyphosphates increase [Ca²⁺]_i in rat MC, in a similar manner to ATP or Ang II and lead to a contraction of MC, suggesting that these nucleotides are also involved in the control of glomerular haemodynamics.     

Influence of microtubules on vascular smooth muscle contraction

Microtubules are ubiquitous in eukaryotic cells and play key roles in many cellular activities. The purpose of this study was to investigate the influence of microtubules on vascular smooth muscle contraction. Quantitative immunocytochemical analysis of rat aortic tissue revealed that, relative to the control group, colchicine (15 M, 90 min) and nocodazole (15 M, 90 min) decreased the microtubule density by 40–50% while taxol (10 M, 90 min) increased the microtubule density by 33%. Isometric contraction studies demonstrated that both colchicine and nocodazole caused an upward shift in the phenylephrine (10^–8 to 10^–5 M) dose–response curve while taxol caused no significant change when compared to the control group. Potassium chloride (30 mM) induced 55 ± 5% P ₀ contraction in DMSO treated vessel rings. The active tension increased to 73 ± 5% P ₀ and 71 ± 6% P ₀ after pretreatment of the aortic rings with colchicine or nocodazole, respectively. Taxol did not cause a significant change in the active tension (56 ± 7% P ₀). These results indicate that microtubule depolymerization enhances isometric contraction of vascular smooth muscle and this enhanced contraction is not receptor dependent. Pretreatment of the aortic rings with an inhibitor of nitric oxide synthase (NOS) (N -nitro-L-arginine) did not change the increased contractile response to phenylephrine due to microtubule depolymerization suggesting that this phenomenon is not mediated by endothelium dependent relaxation.     

Effect of depolarizing and hyperpolarizing agents on the membrane potential difference of primary cultures of rabbit aorta vascular smooth muscle cells

Vascular smooth muscle cells of rabbit aorta were enzymatically dispersed, kept in primary culture, and studied between days 1 and 7 in a bath rinsed with Ringer-like solution at 37°C. The electrical membrane potential difference (PD) was measured with microelectrodes. The mean value of PD was –50±0.4 mV (n=53). Cromakalim (BRL 34915), 1 mol/l and 10 mol/l, hyperpolarized the membrane potential by 9±1 mV (n=11) and 15±1 mV (n=53) respectively. Glibenclamide (10 mol/l) abolished the hyperpolarizing effect of cromakalim (n=6). Simultaneous addition of cromakalim and glibenclamide (both 10 mol/l, n=11) and glibenclamide itself (10 mol/l, n=7) had no effect on PD. In patch-clamp experiments in outside-out-oriented Ca²⁺-sensitive K⁺ channels, cromakalim increased the open probability (P _o) only slightly and only with a cytosolic Ca²⁺ activity of 1 mol/l. In all other series cromakalim had no effect on the P _o of these channels. Forskolin (10 mol/l) hyperpolarized PD by 6±1 mV (n=13). The nucleotides UTP, ATP and ITP (10 mol/l) depolarized PD by 12±1 mV (n=7), 8±1 mV (n=65) and 5±1 mV (n=6) respectively. GTP, [,-methylene]ATP and adenosine had no significant effect. Mn²⁺ (1 mmol/l, n=18), Ni²⁺ (1 mmol/l, n=13), Co²⁺ (1 mmol/l, n=11), Zn²⁺ (1 mmol/l, n=6) and the Ca²⁺-channel blockers verapamil and nifedipine (both 0.1 mmol/l, n=6) did not attenuate the depolarization induced by 10 mol/l ATP. Fetal calf serum (100 ml/l, n=7) depolarized PD by 11±2 mV. This effect was not abolished by nifedipine or by replacing NaCl by choline chloride. The data indicate that PD of vascular smooth muscle cells is depolarized by P₂ agonists and hyperpolarized by the K⁺-channel opener cromakalim. The effect of cromakalim is antagonized by glibenclamide. The effect of cromakalim is probably not mediated by the K⁺ channel identified in excised patches.Supported by DFG Gr 480/10     

Transient improvement of polymorphonuclear leukocyte function by splenectomy in β-thalassemia

Host defense mechanisms in transfusion-dependent non-splenectomized patients with -thalassemia were studied. Polymorphonuclear leukocytes (PMNLs) of non-splenectomized patients responded poorly to zymosan generated chemotactic factors. Chemotactic indices were 22.1 m ± 2.8 (mean ± S.D.) using zymosan activated serum (ZAS) as the attractant in comparison to 20.4 m ± 2.6 when fresh untreated serum was used. In contrast, chemotactic indices of normal PMNLs increased from 21.1 m to 33.6 m ± 3.1 in response to ZAS. Normal PMNL responses to a mixture of normal ZAS and thalassemic serum were inhibited; the mean chemotactic index was 18.1 m ± 5.1 with use of ZAS alone. Splenectomy temporarily reverses these alterations. Adherence to nylon wool of PMNLs suspended in fresh thalassemic serum prior to splenectomy was 3.1% ± 1.1 (mean ± S.D.); 20 days after splenectomy adherence increased to 14.0% ± 2.8 (P = 0.0001) and remained at this level for 90 days. At 120 and 150 days after splenectomy adherence decreased to 1.5% ± 0.8 and 1.0% ± 0.85 respectively. Splenectomy also transiently abrogated the failure of zymosan to generate chemotactic factors in thalassemic serum.This study was presented in part at the American Federation for Clinical Research, Central Society, Infectious Diseases, Chicago, Illinois, November 3, 1983     

Life cycle ofHepatozoon mehlhorni. sp. nov. in the viperEchis carinatus and the mosquitoCulex pipiens

Hepatozoon mehlhori sp. nov. and its developmental stages from the tissues of the Egyptian viperEchis carinatus and the mosquitoCulex pipines are described. The erythrocytic parasites were differentiated into the small form (trophozoite) measuring 14.5±0.6×4±0.12 m and the mature form (gametocyte) measuring 17.2±1.6×5.4±0.5m. Merogony took place in the pulmonary endothelial cells and in the parenchyma cells of the liver and spleen of the infected vipers. Two types of meront were found. The large meronts (macromeronts) were 30.2±1.73×22.6±1.2 m in size and yielded 16–40 (average 28) micromerozoites measuring 17.2±0.7×5±0.15 m. The small, meronts (micromeronts) measured 18.2±0.6×13.5±0.5 m and yielded 2–14 (average, 8) macromerozoites that were 15.1±0.12×6.2±0.8 m in size. After syzygy in the haemocoel of the mosquito, the microgamont produced four uniflagel-late microgametes (6.4±0.3×4.5±0.5 m in size, with a short flagellum measuring 3.2±0.1 m); on the 3rd day post-infection (p.i.)., one of these fertilized the macrogamete, giving rise to the zygote. The oocyst developed from the zygote on the 5th day p.i. and measured 135±2.6×120±1.8 m. About 11–60 (average, 35) sporoblasts were formed by centripetal invaginations from each oocyst on the 8th day p.i. and developed into sporocysts on the 14th day p.i. Inside each sporocyst, 5–12 (average, 8) sporozoites, each measuring 12.6±1.2×4.1±0.3 m, developed on the 16th day p.i. According to the above-mentioned characteristics the parasite was recorded as being a new species and was namedHepatozoon mehlhorni. Experimental transmission was accomplished by i.p. inoculation of the infectious stages (sporozoites) into uninfected vipers and led to the appearance of blood stages at 4–6 weeks p.i.Abbreviations BLC Blood capillary - DMS developing merozoites - DSP developing sporoblasts - E erythrocyte - F flagellum - HC host cell - HN host nucleus - M Meront - MA macrogamont/macrogamete - MG Microgamete - MIG microgamont - MS merozoites - N nucleus - NG micleus of the microgamete - OC oocyst - P erythrocytic parasite - PV Parasitophorous vacuole - SP sporoblast (s) - SPC sporocyst (s) - SPR sporozoite (s) - ZY zygote (young oocyst)     

A pharmacological study of the control of nasal cooling in the dog

Summary Clearance studies were performed in order to examine the effect of expansion of extracellular fluid volume (ECFV) on the maximal reabsorptive capacity for inorganic phosphate (Tm_Pi) in acutely parathyroidectomized (PTX) and intact rats. Tm_Pi values were obtained in both control and volume expansion. In PTX rats, the Tm_Pi values in control and expansion were 8.25±1.52 and 6.14±1.02 mol/min (mean values ±S.D.), respectively; the Tm_Pi/ GFR values were 2.96±0.31 and 2.09±0.30 mol/ml, respectively. Inintact rats, the Tm_Pi values in control and expansion were 3.56±0.94 and 2.98 ±0.94 mol/min, respectively, and the Tm_Pi/GFR values were 1.34±0.23 and 1.05±0.23 mol/ml, respectively. From these results it is concluded that expansion of ECFV decreases the Tm_Pi values both in the absence and presence of parathyroid hormone.This work was supported in part by a grant from the Deutsche Forschungsgemeinschaft (Fr 239/5)     

Suppressed thyroid-stimulating hormone secretion in patients treated with interleukin-2 and interferon-α2b for metastatic melanoma

Recent reports suggest that combined therapy with recombinant interleukin (IL)-2 and interferon (IFN) alb may result in autoimmune-induced thyroid dysfunction. We prospectively analyzed thyroid function for 6 weeks in two groups of patients with progressive metastatic melanoma treated according to two different protocols. In group I (n =17) three treatment cycles were given, each with three weeks of subcutanous administration of rIL-2 and INF-_2b at different doses. In group 11 (n=13) the chemotherapeutic agent dacarbazine was given in addition. In group 1 three patients developed frank hyperthyroidism, which required antithyroid drug therapy in one case. Autoantibodies against thyroid microsomal antigen, thyroglobulin, and the thyroid-stimulating hormone (TSH) receptor were not significantly elevated in any of these patients. However, the remaining 14 patients showed a significant decrease in TSH after 6 weeks of treatment, from 1.8 ± 0.9 to 0.7 ± 0.7 U/ml (P < 0.02). Thyroid hormones (triiodothyronine, thyroxine, free thyroxine) also increased during the observation time, but this did not parallel the drop in TSH levels. Only thyroxine increased above the upper limit of normal, while triiodothyronine and free thyroxine stayed within the normal range. In group 11, 6 of 13 patients (46%) had a decreased TSH after 6 weeks of treatment. Mean TSH was 1.5±1.4 before and 0.8 ± 0.6 U/ml after 6 weeks and was totally suppressed in three cases. None of these patients showed ouvert hyperthyroidism. Hypothyroidism was not observed in either group. We conclude that treatment with rIL-2 and INF-_2b may not only be associated with autoimmune thyroiditis and hyperthyroidism but also results in suppression of TSH levels while the patients remain euthyroid.Abbreviations IL interleukin - INF interferon - TSH thyroid-stimulating hormone - T3 triiodothyronine - T₄ thyroxine - fro free thyroxine Correspondence to: H. Mönig     

Calcium sensitivity and unloaded shortening velocity of hypertrophied and non-hypertrophied skinned human atrial fibres

The mechanical properties of myocardium of different animals are modified by a chronic increase in haemodynamic load. In this study differences in calcium sensitivity and maximum unloaded shortening velocity of hypertrophic and non-hypertrophic chemically skinned human atrial fibres are characterized. Investigating right atria of 34 patients, possible correlations are studied between preoperative atrial pressure, degree of hypertrophy (estimated from the muscle fibre diameter), calcium responsiveness (pCa₅₀ eliciting half-maximum contraction) and V _max (unloaded shortening velocity). Hypertrophic fibres from atrial appendages of patients having an increased right atrial pressure (RAP 8.5±1.6 mm Hg) and suffering from mitral valve disease (stenosis and insufficiency combined) had a fibre diameter of 18.0±0.9 m. They also had a higher calcium sensitivity (pCa₅₀ 5.65±0.08) and a lower unloaded shortening velocity (1.7±0.1 muscle lengths/s) than non-hypertrophic fibres from the appendages of patients with normal right atrial pressure (RAP 3.2±0.5 mm Hg) and coronary heart disease (CHD: pCa₅₀ 5.45±0.04; V _max= 3.4±0.2 muscle lengths/s; fibre diameter 12.8±0.4 m). Thus non-hypertrophic fibres from control CHD patients differed significantly (p < 0.01) from hypertrophied atrial fibres of patients with mitral valve disease and with combined valve disease (MAV, pCa₅₀=5.58±0.05, V _max 2.0±0.3 muscle lengths/s, fibre diameter 14.6±0.9 m) or aortic valve disease (stenosis combined with insufficiency, fibre diameter 14.8±1.4 m, pCa₅₀ 5.56±0.03, V _max 2.0±0.24 muscle lengths/s; RAP 11.0±2.6 mm Hg). Such alterations of calcium responsiveness, shortening velocity and fibre thickness may reflect an adaptation to the chronic overload in atria from patients with various forms of heart valve disease.     

Parallel changes in red blood cell and renal Na−K-ATPase activity in adrenal and electrolyte disorders in the rat

To compare the activity of Na–K-ATPase in the red blood cells (RBCs) and in renal tissue in disorders of Na⁺ metabolism, the following groups of rats were studied: 1) control, intact rats, 2) adrenalectomized (ADX) rats, 3) intact rats treated with DOCA, 4) ADX DOCA-treated rats, 5) intact salt-loaded rats, 6) ADX salt-loaded rats, 7) intact dexamethasone-treated rats (DEXA), and 8) ADX DEXA-treated rats. After adrenalectomy (group 2) serum Na¹ decreased and serum K⁺ increased.Renal Na–K-ATPase in cortex, medulla and papilla of the control group was 44±2.7 mol Pi/mg prot/h, 128.2±5.9 and 44±3.2 respectively and in group 2 the enzyme activity was 32.5±2.0 (P<0.005), 81.7±4.5 (P<0.001) and 23.6±1.9 (P<0.001) respectively. RBCs Na–K-ATPase of control animals was 2.82±0.19 mol Pi/mg prot/h, while in group 2 the activity was 1.43±0.24 (P<0.001). DOCA treatment of ADX rats (group 4) normalized serum electrolytes and Na–K-ATPase activity in the renal cortex and papilla and in the RBCs. In the renal medulla the correction by DOCA was only partial. Salt loading of ADX rats (group 6) normalized serum electrolytes and Na–K-ATPase activity in the renal medulla and RBCs. Salt loading of normal rats increased RBC Na–K-ATPase to 3.72±0.36 (P<0.02) and medullary Na–K-ATPase to 185.6±9.8 (P<0.01). DEXA treatment of ADX rats (group 8) corrected only partially the abnormalities in serum electrolytes and Na–K-ATPase activity in the kidney and in the RBCs. These findings show, 1) parallel changes in the activity of Na–K-ATPase in the RBCs and in the kidney after adrenalectomy, 2) parallel changes in the enzyme activity in RBCs and in the kidney medulla after salt loading, and 3) correction towards normal of RBC Na–K-ATPase after ADX by NaCl and DOCA treatment.This study was supported by the Morton S. Kaufman Hemodialysis Foundation     

Renal function in normal and potassium-depleted rats before and after preparation for micropuncture experimentation

Renal function was examined in unrestrained conscious rats maintained on either a control diet or a low-potassium diet, then re-examined in the same animals after thiobutabarbital (Inactin) anaesthesia and preparation for micropuncture studies. In conscious rats, inulin clearance (C_In) was not significantly different in the two groups (control 1012±43, low-K 904±58 l/min per 100g body wt; mean±SE), but lithium clearance (C _Li; used as an estimate of end-proximal fluid delivery) and fractional lithium excretion (FE_Li) were substantially reduced in the low-K group (C _Li: 246±11 vs 126±8 l/min per 100g body wt, P<0.001; FE_Li: 0.245±0.009 vs 0.143±0.008, P<0.001). Following anaesthesia and preparation for micropuncture, there were significant reductions in urine flow rate and sodium excretion in the control group, but not in the low-K rats. Potassium excretion increased in both groups, but values in the potassium-depleted animals remained extremely low. In neither group of rats was preparation for micropuncture associated with significant changes in C _In, C _Li or FE_Li. Thus, differences in tubular function between control and potassium-depleted rats were still apparent. The results suggest that preparation for micropuncture disturbs the function of the distal nephron, but that rates of glomerular filtration and proximal tubular reabsorption remain similar to values in conscious animals.     

PTH independent sex difference in renal handling of inorganic phosphate in the rat: effect of oophorectomy

In order to evaluate the sex difference in the renal handling of inorganic phosphate (Pi) in the rat we performed clearance experiments using intact, thyroparathyroidectomized (TPTX), oophorectomized (OophX) and orchiectomized (OrchX) rats. During stepwise elevation of the Pi concentration in plasma (Pi-titration) to about 6 mmol/l the reabsorptive mechanism of Pi was saturated. The ratio Pi-reabs./GFR in intact males was higher than in females. A significant difference in this parameter was also observed in thyroparathyroidectomized rats: in females this value was 3.47±0.13, and in males it was 4.54±0.37 mol/ml (P<0.001). Oophorectomy in the absence of parathyroid hormone (PTH) increased Pi-reabs./GFR from 3.18±0.36 to 4.12±0.24 mol/ml (P<0.001); however, orchiectomy did not significantly change the reabsorption of Pi. In conclusion, the present results demonstrate a PTH independent sex difference in the renal handling of inorganic phosphate and are consistent with the hypothesis that estrogens may play a dominant role in this differentiation.This work was supported by a grant from the Deutsche Forschungsgemeinschaft (Fr 239/9-1)Dedicated to Professor Dr. Karl Julius Ullrich     

Increase of cytochromes a and a3 in rat kidney mitochondria in response to administration of aldosterone in vivo

Summary The influence of aldosterone in vivo on cytochromes in rat kidney mitochondria is studied, comparing rats in normal state, adrenalectomized state, and adrenalectomized rats 2 hours after administration of adlosterone (7.5 g per 100 g rat as single dose plus infusion of 0.125 g per hour and 100 g rat).No significant changes between these states are observed of either cytochrome b, cytochrome c₁, or cytochrome c content.Cytochrome a, as estimated from the 605 m band of the difference spectrum (reduced vs. oxidized), decreases from 508±51 nanomoles per g protein (n=12) in the controls to 273±65 (n=12) nanomoles per g protein in the adrenalectomized state.After administration of aldosterone to adrenalectomized rats the concentration of cytochrome a increased to 464±38 nanomoles per g protein (n=12).The difference spectrum (anaerob vs aerob+Antimycin A) shows that the absorbancy maximum at 444 m (more specific for cytochrome a₃) is also considerably decreased after adrenalectomy, and increased after administration of aldosterone to adrenalectomized rats. Hence the contents of both compounds of cytochrome oxidase, cytochrome a and cytochrome a₃, appear to be controlled by aldosterone.The molar ratio of cytochrome c to cytochrome a is 1.1 in the controls, 1.8 after adrenalectomy, and 1.1 after administration of aldosterone to adrenalectomized rats.Possible relations of this effect to the previously observed increase of tricarboxylic acid cycle enzyme activities under aldosterone are discussed.The 46% decrease of the cytochrome a content in mitochondria from the whole kidney of adrenalectomized rats, which is restored to the normal level in response to aldosterone, is difficult to reconcile with the concept that the hormone acts on distal tubules only, as these constitute only about 10% of the material used. Therefore, the present result is considered to support the concept that the hormone acts on both distal and proximal tubules.     

Changes in sarcoplasmic metabolite concentrations and pH associated with the catch contraction and relaxation of the anterior byssus retractor muscle ofMytilus edulis measured by phosphorus-31 nuclear magnetic resonance

Summary The sarcoplasmic concentrations of phosphorus metabolites and pH (pH_in) were measured in the anterior byssus retractor muscle (ABRM) ofMytilus edulis by³¹P nuclear magnetic resonance spectroscopy. During an active contraction induced by 10^–3 m acetylcholine, the concentration of arginine phosphate ([Arg-P]_in) decreased from the resting value of 7.47±0.26 (mean±se,n=8) to 6.67±0.29 (n=6) mol g^–1, and that of inorganic phosphate (P_i) consistently increased from 0.84±0.06 (n=7) to 1.61±0.12 (n=5) mol g^–1. In the catch state following the active contraction, these concentrations were close to their resting levels, indicating that the catch is an inactive state. 5-hydroxytryptamine caused a rapid relaxation of the catch, which was associated with a slight decrease in [Arg-P]_in and an increase in pH_in by ca 0.2 units. The sarcoplasmic concentration of ATP (mean, 1.6mol g^–1) did not change throughout the contraction-relaxation cycle.