Morphine Stimulates Mesangial Cell TNF-α and Nitrite Production

Background: Intravenous opiate abusers are susceptible to develop heroin and HIV-associated nephropathies; however, the role of opiates in the development of these kidney lesions is not clear. Patients with opiate addiction are prone to recurrent infections. Methods: The effect of morphine was studied on the generation of TNF- with or without LPS (lipopolysaccharide) by cultured mouse mesangial cells. In addition, the effect of morphine was evaluated on mesangial cell nitrite production. To evaluate the role of opiate receptors, we studied the effect of naloxone and naltrexone on mesangial cell TNF- and nitrite production. To determine the role of TNF- on mesangial cell nitrite production, we examined the effect of anti-TNF- antibody on morphine-induced nitrite production. Assay of TNF- and nitrite production was carried by ELISA and Griess method respectively. Results: Morphine alone did not enhance the generation of TNF- by mesangial cells, however, an enhanced (P < 0.001) TNF- production was observed when mesangial cells were first treated with morphine for 18 h and then activated further with LPS. Maximum release of TNF- was seen at a concentration of 10^–12 M of morphine. Opiate receptor antagonists (naloxone and naltrexone) inhibited the effect of morphine. Morphine also amplified (P < 0.0002) the effect of LPS on mesangial cell nitrite production. Anti-TNF- antibody attenuated morphine induced nitrite generation. Conclusion: We conclude that morphine stimulates the generation of TNF- by LPS-activated mesangial cells. This effect of morphine seems to be opiate receptor mediated and has a downstream effect in the form of mesangial cell nitrite generation. The present in vitro study provides the basis for a hypothesis that morphine may be playing a role in the development of heroin and HIV-associated nephropathies.     

The tubular vacuolation process in amphibian skeletal muscle

The exposure of amphibian muscle to osmotic shock through the introduction and subsequent withdrawal of extracellular glycerol causes vacuolation in the transverse tubules. Such manoeuvres can also electrically isolate the transverse tubules from the surface (detubulation), particularly if followed by exposures to high extracellular [Ca2+] and/or gradual cooling. This study explored factors influencing vacuolation in Rana temporaria sartorius muscle. Vacuole formation was detected using phase contrast microscopy and through the trapping or otherwise of lissamine rhodamine dye fluorescence within such vacuoles. The preparations were also examined using electron microscopy, for penetration into the transverse tubules and tubular vacuoles of extracellular horseradish peroxidase introduced following the osmotic procedures. These comparisons distinguished for the first time two types of vacuole, open and closed, whose lumina were respectively continuous with or detached from the remaining extracellular space. The vacuoles formed close to and between the Z-lines, but subsequently elongated along the longitudinal axis of the muscle fibres. This suggested an involvement of tubular membrane material; the latter appeared particularly concentrated around such Z-lines in the electron-micrograph stereopairs of thick longitudinal sections. Open vacuoles formed following osmotic shock produced by extracellular glycerol withdrawal from a glycerol-loaded fibre at a stage when one would expect a net water entry to the intracellular space. This suggests that vacuole formation requires active fluid transport into the tubular lumina in response to fibre swelling. Closed vacuoles only formed when the muscle was subsequently exposed to high extracellular [Ca2+] and/or gradual cooling following the initial osmotic shock. Their densities were similar to those shown by open vacuoles in preparations not so treated, suggesting that both vacuole types resulted from a single process initiated by glycerol withdrawal. However, vacuole closure took place well after formation of open vacuoles, over 25 min after glycerol withdrawal. Its time course closely paralleled the development of detubulation reported recently. It was irreversible, in contrast to the reversibility of open vacuole formation. These findings identify electrophysiological detubulation of striated muscle with closure of initially open vacuoles. The reversible formation of open vacuoles is compatible with some normal membrane responses to some physiological stresses such as fatigue, whereas irreversible formation of closed vacuoles might only be expected in pathological situations as in dystrophic muscle.This revised version was published online in September 2005 with corrections to the Cover Date.     

Kombinationsformen der Arteriitis

Zusammenfassung Ausgehend von den UntersuchungenRössles zum Formenkreis der rheumatischen Gewebsveränderungen, wird an Hand von 4 eigenen Fällen die Problematik der morphologischen Differentialdiagnose zwischen rheumatischer Arteriitis, Periarteriitis (Panangiitis) nodosa und Thrombangitis obliterans unter Berücksichtigung der Literatur aufgezeigt. Mit der morphologischen Definition der verwendeten Begriffe fibrinöse Entzündung und proliferierende und granulierende Angiitis wird versucht, die drei Formen der Gefäßwandentzündung morphologisch abzugrenzen. Die überschneidungen der morphologischen Symptome und die Kombinationsformen werden aus Beschreibung und Abbildung einschlägiger veröffentlichter Fälle aufgezeigt. Aus dem Nachweis dieses Schrifttumsvergleichs und dem der Differentialdiagnose der 4 eigenen Fälle wird gefolgert, daß bei solchen nicht-klassischen Gefäßentzündungen die Annahme von Kombinationsformen richtiger ist als die immer stärkere Erweiterung des sog. rheumatischen Formenkreises.

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Summary With reference to the investigations ofRössle On the Classification of the Rheumatic Tissue Changes, the problems that are associated with the differential diagnosis of rheumatic arteritis, periarteritis (panangiitis) nodosa, and thromboangiitis obliterans are discussed, exemplified by the report of four cases, and by a review of the literature. By applying the morphological criteria of fibrinous inflammation, proliferative and granulomatous angiitis, an attempt is made histologically to differentiate these three forms of inflammation of the vascular wall. The overlapping of the morphological criteria and the combined forms that mav occur are illustrated in the four cases reported. As shown by comparing the cases in the literature, and as exemplified by the differential diagnosis of the four cases, the inference is made, that with such types of non-classical vascular inflammation, it is better to refer to combined forms than to the more diffuse so-called rheumatic classification.

     

Cytokine mRNA in the Joints and Draining Lymph Nodes of Rats with Adjuvant Arthritis and Effects of Cyclosporin A

TNF- and IL-1 promote leukocyte recruitment to arthritic joints and may contribute to cartilage degradation while regulatory cytokines such as IL-4 and IL-1RA may in part determine the course of arthritis. Here we report the pattern of TNF-, IL-1, IL-6, IFN-, IL-1RA, and IL-4 mRNA expression, detected by RT/PCR, in the talar joint and draining popliteal lymph node (PLN) of rats with adjuvant arthritis (AA). Levels of TNF- and IFN- mRNA were increased in the PLN before clinical signs of arthritis. This was followed by increases in IL-1 and IL-1RA mRNA at d9 and IL-6 mRNA at d12. PLN IL-1RA mRNA levels were positively correlated with those of IL-1 and TNF- throughout d5-d20. IL-4 mRNA levels were highest on days 7 and 20. In the synovium, a small increase in TNF-, IL-1, and IL-6 mRNA was detected on d5 then again on d12. Maximal synovial TNF- levels were reached on d20, while IL-1 peak expression was on d16 and IL-6 on d14. IL-4, IL-1RA, and IFN- mRNA was undetectable in the synovium. Cyclosporin treatment for 4 days, initiated at the height of arthritis, rapidly decreased clinical disease, and decreased migration of neutrophils and T lymphocytes into the joints. Yet no significant effect of CyA was observed on inflammatory cytokine expression, although the correlation between PLN IL-1RA and IL-1 or TNF- was lost in treated animals. Thus there is a variable pattern of cytokine gene expression in rat AA, the undetectable IL-4 and IFN- mRNA in synovium being analogous to human rheumatoid arthritis.     

Characterization and localization of α-connectin (titin 1): An elastic protein isolated from rabbit skeletal muscle

Summary A simplified procedure to isolate-connectin (titin 1, TI), a gigantic elastic protein, from rabbit skeletal muscle is described. A rapid column chromatography step to concentrate-connectin is introduced. Separation of-connectin from-connectin is introduced. Separation of-connectin from-connectin (titin 2, TII) in the presence of 4 M urea at pH 7.0 did not cause any change in the secondary structure of-connectin as judged by circular dichroic spectra. Ultraviolet absorption spectra and the amino acid composition of-connectin (MW, approximately 3×10⁶) were similar to those of its proteolytic product,-connectin (MW, approximately 2×10⁶). Circular dichroic spectra suggested that both- and-connectin consist of 60%-sheet and 30%-turn. It thus appears that the whole elastic filament of connectin has a folded-strand structure. Proteolysis of-connectin by calpain resulted in formation of-connectin and smaller peptides. The-connectin interacted with both myosin and actin filaments similarly to-connectin. Polyclonal antibodies raised against 1200 kDa peptides obtained from aged rabbit skeletal myofibrils reacted with-connectin (titin 1, TI) but only weakly with-connectin (titin 2, TII) in rabbit skeletal muscle. Immunoelectron microscopy and indirect immunofluorescence microscopy revealed that the antibodies bound at the Z-line and at the epitope regions in the I-band near the binding site of a monoclonal antibody SMI whose position depends on sarcomere length. It thus appears that-connectin extends from the edge of M-line to the above epitope region in the I-band.     

The effects on primary and secondary spindle afferents of fusimotor reflex during ramp and sinusoidal muscle stretch

Summary Tetanic stimulation of ipsilateral Triceps surae afferents leads to a reflex increase in the initial burst from the primary spindle afferents during ramp stretch in both intercollicular decerebrate and spinal preparations. The dynamic index does not change significantly.Slow, low-amplitude sinusoidal muscle stretches (1.6 Hz, 0.55 mm peak-peak) disclose an increased dynamic sensitivity of primary afferents during reflex activation. The same activation in the statically adapted muscle results in a great many cases in little or no increase in spindle discharge rate.In the frequency-stretch relation, the discharge rates of primary afferents in the spinal cat and of secondaries in decerebrate animals are greatly increased by combined fusimotor activation and low-amplitude sinusoidal stretch.We conclude that both static and dynamic motoneurons are implied in this reflex.     

Development of pump electrogenesis in hypokalaemic rat muscle

In hypokalaemic rats maintained on a potassium deficient diets for 10–50 days, the isolated Na-loaded and K-depleted (Na-rich) muscle fibers showed the membrane potential less than –115 mV in fresh muscles of normal rats in K⁺-free Krebs solution. Upon adding 5 mM K⁺ to the K⁺-free medium bathing the soleus muscles, the measured potentials of Na-rich muscles always exceeded the membrane potentials of fresh muscles in 5 mM K⁺. The hyperpolarization was dependent on the amount of intracellular Na⁺ concentration ([Na]_i) accumulated during the potassium deficiency. The electrogenic Na-pump was activated by an increase of [Na]_i of less than 5 mM. Further increases in [Na]_i resulted in increases in membrane potential which appeared to approach a limit at [Na]_i levels higher than 65 mM.     

Prostate specific antigen and prostate specific acid phosphatase in adenocarcinoma of Skene's paraurethral glands and ducts

An autopsy case of adenocarcinoma of Skene's paraurethral gland co-incident with renal cell carcinoma is described. The adenocarcinoma showed distinct prostate specific antigen and prostate specific acid phosphatase pointing to the equivalence between the male prostate and Skene's paraurethral glands and ducts. Skene's gland are the homologue of the prostate in females and tumours arising from them are immunohistochemically similar to male prostate carcinoma.In the title and text the authors used the official term of Nomina Anatomica paraurethral (Skene's) glands and ducts. Nevertheless recently published data on cross-antigenicity between the male prostate and Skene's glands and the newly discovered exocrine and neuroendocrine parameters of the prostate homologue in the female, comparable with the male prostate (Zaviai 1987), support the use of the same term — the prostate — for prostatic tissue in both sexes (Zaviai 1987, Zaviai et al. 1985). The designations female prostate homologue or female prostate equivalent are a compromise between terms the female prostate and Skene's paraurethral glands.     

Bioactive 5-Reduced 16α,17α-Cyclohexanoprogesterone Derivatives Weakly Interact with Proteins of the Soluble Uterine Fraction

We studied competitive activities of 16,17-cyclohexano-5- and 5-dihydroprogesterone in replacing ₃H-progesterone and ₃H-16,17-cyclohexano-6-methylprogesterone from protein complexes. Direct binding of ₃H-5-reduced derivatives with proteins of soluble fractions from rat and rabbit uteri was also assayed. Cd values for 5-reduced derivatives were in the micro- or submicromolar range. The data suggest that biological effects of these analogues are not mediated via soluble uterine receptors.     

Differential effects of interleukin-1α and β on the arachidonic acid cascade in human synovial cells and chondrocytes in culture

The effects of interleukin-1 and were tested on the [³H]-arachidonic acid release and the prostaglandin synthesis by human cultured synovial cells and chondrocytes. Both forms of interleukin-1 stimulated the arachidonic acid release but interleukin-1 was more potent than IL-1. Human synovial cells and chondrocytes synthesized three types of prostaglandins upon stimulation with interleukin-1 or : prostaglandin E₂, F₂ and 6-keto-prostaglandin F₁. Regarding the synthesis of these prostaglandins, IL-1 was again more potent than IL-1. A comparison between interleukin-1-stimulated synovial cells and chondrocytes revealed neither significant quantitative nor qualitative differences in both the arachidonic acid release and the prostaglandin synthesis.     

Presynaptic excitability changes induced by morphine in single cutaneous afferent C- and A-fibers

Intraspinal microstimulation was used to test the excitability of single cutaneous (sural) A- and C-fibers at their central terminals in the cat. Systemic morphine (1.0–2.6 mg/kg) increased the antidromic threshold to 110% (mean value) of control in C-fibers. In some of the C-fibers this effect was reversed by naloxone. A-fibers were not consistently affected by mophine.Preliminary reports were given at the NIMH-conference on Endorphins in mental health research, held in San Juan, Puerto Rico (1977), and at the joint meeting of the British and German Physiological Societies, Cambridge/England (July 1978)     

Involvement of β-adrenoreceptors in the shift of ocular dominance after monocular deprivation

Summary In the visual cortex of young kittens, the extent of ocular dominance shift following brief monocular lid suture was less than expected in the hemisphere which had been locally perfused with -adrenergic receptor blockers at a moderate concentration. The present result was not due to the effect similar to binocular deprivation, since local anesthetic effect of -adrenergic blockers was unlikely to be involved in the present paradigm. Thus, the present results are consistent with the idea that -adrenergic receptors within the visual cortex are necessary, at least in part, for the regulation of visuocortical plasticity.     

The ansa cervicalis and the infrahyoid muscles of the rat

Summary Perikarya of motoneurons and spinal ganglion cells attributed to infrahyoid muscle nerves of the rat were labelled by retrogradely transported horseradish peroxidase (HRP). For the differentiation of motor and sensory axons cross sections of the nerves were stained for acetylcholinesterase. Numbers and diameter distributions of perikarya and myelinated axons were determined.Motoneuronal perikarya innervating the infrahyoid muscles are located from the transition zone brain stem/spinal cord to the segment C 3. They are found mostly in the medial part of the Rexed laminae VII and VIII at the level of C 1 and C 2 and more ventrolaterally in C 3 and are therefore located to a large extent in areas until now not recognized to contain motoneurons. Our results provide evidence for a somatotopic organization of the motoneurons in the upper cervical spinal cord.The diameter distributions of motoneuronal perikarya and axons are in most cases bimodal, the two modes corresponding to -and -motoneurons. In relation to the diameters of their perikarya -axons are significantly thicker than -axons. In contrast to the motoneurons no clear correlation could be established between the sizes of perikarya of spinal ganglion cells and their peripheral processes.This work was partly supported by the Hartmann Müller-Stiftung Zürich     

Untersuchungen zur Ultrastruktur lympho-epithelialer Thymustumoren unter besonderer Berücksichtigung der sog. „Emperipolesis“

Summary The ultrastructure of eleven thymomas with lymphocytic predominance, one epitheloid cell thymoma and two normal human thymuses is described with special reference to Emperipolesis. All patients have had myasthenia gravis.The normal human thymus consists of three parts: outer cortex, inner cortex, and medulla. The outer cortex contains mainly lymphoblasts and Metcalf's macrophages within the so-called Clark-packet's. The inner cortex consists mainly lymphocytes and interdigitating reticulum cells, and the medulla of epithelial cells, lymphocytes and Hassall's corpuscles.In all cases of lympho-epithelial thymoma and in normal human thymuses there are enormous interdigitations between epithelial (tumor) cells, lymphocytes and macrophages. The epitheloid cell thymomas also show findings which suggest an epithelial cell interaction. We have not found intact lymphocytes inside the cytoplasm of normal and/or tumor epithelial cells, macrophages or interdigitating reticulum cells.The intracellular existence of intact lymphocytes has been termed Emperipolesis by Humble, Jayne, and Pulvertaft, meaning internal wandering. These investigations indicate that Emperipolesis is not an adequate term for cellular interaction in normal human thymuses and thymomas. A false impression of intraepithelial location of thymic lymphocytes is created by two-dimensional sections of complex thymic structure. These ultrastructural studies revealed damage to lymphocytes only in macrophages with lymphocytolysis within these cells and accumulation of numerous heterophagic vacuoles containing fragments of lymphocytic debris within them.

FrÄulein C. Schürmann danke ich für die gute technische Assistenz, Herrn Priv.-Doz. Dr. med. R. W. Ch. Janzen, Neurologische Klinik der UniversitÄt Hamburg, für die klinischen Daten der Myasthenie-Patienten     

Effect of BW443C81, a novel opioid,on non-cholinergic bronchoconstrictor responses and neurogenic plasma extravasation in the guinea pig

The novel, peripherally acting opioid peptide, BW443C81, which attenuates airway sensory nerve impulses, was examined on non-cholinergic (NC) constrictor responsesin vitro andin vivo and neurogenic plasma extravasationin vivo in guinea-pig airways. Non-cholinergic contractions of guinea pig isolated bronchi, evoked by electrical field stimulation, were concentration-dependently inhibited by BW443C81 and morphine (10 nmol/l-100 mol/l). In anaesthetised, artificially ventilated guinea pigs, frequency-related NC bronchoconstictor responses evoked by antidromic electrical stimulation of the vagus nerves were reduced by BW443C81 (100 g/kg/min i.v. infusion) and morphine (1 mg/kg i.v.). Neurogenic plasma extravasation produced by bilateral electrical vagal nerve stimulation in spontaneously breathing, anaesthetised guinea pigs was also inhibited by BW443C81 (1 mg/kg i.v.). The inhibitory effects of BW443C81 were reversed/prevented by naloxone. BW443C81 inhibits NC bronchoconstrictor responses and neurogenic plasma extravasation in guinea pig airways, consistent with its previously described -opioid receptor-mediated inhibitory action on airway sensory nerve function.     

Intrinsic membrane properties causing a bistable behaviour of α-motoneurones

Summary In decerebrate cats a train of impulses in Ia afferents may lead to a sustained increase in excitability of -motoneurones of homonymous and heteronymous muscles. It was previously suggested that this long-lasting excitability increase reflects a maintained synaptic input to the motoneurones from excitatory interneurones. With intracellular recording from motoneurones we here demonstrate that the sustained increase of -motoneurone activity is due to an all-or-none plateau depolarization. This plateau can be induced by a short train of excitatory synaptic potentials or a brief, intracellularly injected depolarizing current pulse and is terminated by a short train of inhibitory synaptic potentials or a hyperpolarizing current pulse. It is concluded that maintained motor unit firing triggered by a brief train of impulses in Ia afferent reflects an intrinsic bistable behaviour of -motoneurones.     

Bulbar neurones with axonal projections to the trigeminal motor nucleus in the cat

Summary The location of bulbar neurones with axons projecting to the ipsi- and contralateral trigeminal motor nucleus were investigated in cats anaesthetized with sodium pentobarbital. Wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) was injected in amounts of 5–24 nl. A volume-calibrated microelectrode was used for recording of evoked potentials and pressure injection of WGA-HRP. The injection site was guided by the position where a maximal antidromic response was evoked by electrical stimulation of the masseteric nerve. The survival time was 19–22 h. In preparations with the depot located in the masseteric subnucleus retrogradely stained neurones were found bilaterally in the borderzone of the trigeminal motor nucleus. Dense populations of stained neurones were observed ipsi- and contralaterally in the dorsal division of the main sensory trigeminal nucleus and the subnucleus- of the oral nucleus of the spinal trigeminal tract. Clusters of WGA-HRP-neurones were observed bilaterally in the lateral tegmental field at the level of the subnucleus- of the oral nucleus of the spinal trigeminal tract, bilaterally dorsal to the facial nucleus and contralaterally adjacent to the hypoglossal nucleus. No stained neurones were found in the gigantocellular reticular nucleus. A group of stained neurones was located in the marginal nucleus of brachium conjunctivum and some were found in the raphé nuclei near obex. Cell profiles were of two types: medium-sized neurones with a triangular profile and 30–40 m diameter, and fusiform neurones 10×50–70 m. Convergence of descending cortical and trigeminal afferent inputs on interneurones located in the lateral borderzone of the trigeminal motor nucleus, i.e. the intertrigeminal area, is reported in the preceding paper.List of Abbreviations BCM Marginal nucleus of the brachium conjunctivum - CAE Nucleus caeruleus - CI Inferior central nucleus - Cu Cuneate nucleus - Cux External cuneate nucleus - DMV Dorsal motor nucleus of the vagus - FTG Gigantocellular tegmental field - FTL Lateral tegmental field - FTP Paralemniscal tegmental field - Gr Gracile nucleus - Mb Medial borderzone of NVmt - NintV Intertrigeminal area - NsV Supratrigeminal nucleus (area) - NVmes Mesencephalic trigeminal nucleus - NVmt Trigeminal motor nucleus - NVsnpr Main sensory trigeminal nucleus - NVsnpr-d Main sensory trigeminal nucleus, dorsal division - NVsnpr-v Main sensory trigeminal nucleus, ventral division - NVspc Caudal nucleus of the spinal trigeminal tract - NVspo- Subnucleus- of the oral nucleus of the spinal trigeminal tract - NVspo- Subnucleus- of the oral nucleus of the spinal trigeminal tract - NVspo- Subnucleus- of the oral nucleus of the spinal trigeminal tract - V Spinal trigeminal tract - NVII Facial nucleus - VII Facial nerve - NXII Hypoglossal nucleus - XII Hypoglossal nerve - Ols Superior olive - Rb Rostral borderzone of NVmt - Vb Ventral borderzone of NVmt - VIN Inferior vestibular nucleus - VSL Superior vestibular nucleus, lateral division     

Neural nicotinic acetylcholine responses in solitary mammalian retinal ganglion cells

Using the patch-clamp technique,whole-cell recordings from solitary rat retinal ganglion cells in culture have established the nicotinic nature of the acetylcholine responses in these central neurons. Currents produced by acetylcholine (5–20 mol/l) or nicotine (5–20 mol/l) reversed in polarity near –5 mV and were unaffected by atropine (10 mol/l). Agonist-induced currents were blocked by low doses(2–10 mol/l) of the classical ganglionic antagonists hexamethonium and mecamylamine, as well as by d-tubocurarine and dihydro--erythroidine (the latter two do not discriminate clearly between ganglionic and neuromuscular junction receptors). Treatment with the potent neuromuscular blocking agent -bungarotoxin (10 mol/l) did not affect the cholinergic responses of these cells, while toxin F (0.2 mol/l), a neural nicotinic receptor antagonist, readily abolished acetylcholine-induced currents. Thus, the experiments performed to date show that the nicotinic responses of retinal ganglion cells in the central nervous system share the pharmacology of autonomic ganglion cells in the peripheral nervous system. The ionic current carried by the nicotinic channels was selective for cations, similar to that described for nicotinic channels in other tissues. In addition, single-channel currents elicited by acetylcholine were observed in whole-cell recordings with seals > 5 G as well as in occasional outside-out patches of membrane. These acetylcholine-activated events, which had a unitary conductance of 48 pS and a reversal potential of 0 mV, represent the ion channels that mediate the neural nicotinic responses observed in these experiments on retinal ganglion cells.     

Immunological study of HB-Ag carriers

Summary The serum immunoglobulin levels, the presence of circulating autoantibodies, the number of peripheral atypical lymphocytes and the number of DNA and RNA synthesizing mononuclear cells were studied in a group of healthy HB-Ag (australia Antigen) carriers and in a group of healthy HB-Ag negative control subjects.In the HB-Ag carriers, in spite of the persistent antigenaemia, all these immunological findings were normal (as in the control subjects).