A fast perfusion system for single cell physiology optimized for microscopes with water immersion objectives

A perfusion system was constructed which allows the fast application of different solutes underneath a water immersion objective. The perfusion system is mounted into the immersion objective by milling a slot into the frontal metal plate of the lens holder. It consists of a five-channel pipette fixed to the objective and solution reservoirs gated by computer controlled magnetic valves. Up to five different solutions can be applied to the specimen under study. The solution between objective and specimen is completely exchanged after 1–2 s as determined from fluorescence measurements. This arrangement is optimized for [Ca²⁺] measurements with a fluorescence measurement system in tissue slices, where upright microscopes are required. It offers the advantage of saving a micromanipulator for the perfusion pipette and facilitates a fast, reproducible and precise positioning of the perfusion system.     

细胞周期抑制剂P27^Kip1对乳腺癌的预后意义

P2 7Kip1属广谱CDKI家族 ,是一种细胞周期的负向调节剂 ,在细胞增殖、分化、凋亡及肿瘤的发生发展等过程中具有重要的调节作用。人类肿瘤中P2 7基因的改变极为罕见 ,但P2 7蛋白表达的改变与其生物学行为密切相关 ,近年来的诸多研究表明P2 7蛋白表达对包括乳腺癌在内的多种恶性肿瘤具有预后意义     

Substance P modulation of dopamine in the nucleus accumbens

Rats were implanted with bilateral injection cannulae in the nucleus accumbens. While microinjection of substance P (1.0-10.0 micrograms/side) did not alter locomotor or rearing behavior, it did potentiate the motor stimulant effects of intra-accumbens injection with dopamine. Further, substance P injection into the nucleus accumbens significantly elevated the levels of dopamine metabolites in that nucleus. Both the behavioral and neurochemical dose-response curves were biphasic, with 3.0 micrograms of substance P producing a maximal response. These data suggest an interaction between substance P and dopamine in the nucleus accumbens.     

Adriamycin-induced DNA strand breaks in HeLa and in P388 leukaemia cells detected using in situ nick translation

DNA strand breaks produced by adriamycin (ADR) were measured in HeLa cells and ADR-sensitive and -resistant P388 leukaemia cells, using the in situ nick translation method. The break sites in the DNA were translated artificially in the presence of Escherichia coli DNA polymerase I and 3H-labelled dTTP, and were visualized by autoradiographic observation of the grains. The DNA strand breaks in the HeLa cells increased in a dose-dependent manner, compared with findings in the untreated control cells, i.e., 15.2 fold at 20 micrograms/ml of ADR for 1 h. This level correlated with DNA single-strand breaks detected by the alkaline elution method. DNA breaks were also noted in the ADR-sensitive P388 cells, but in the ADR-resistant cells the level of DNA strand breaks was low. The enhanced cytotoxicity is apparently the consequence of the enhanced potential of ADR to cause breaks in the DNA strands. Our findings show that the survival response of the cells decreases and the level of DNA strand breaks increases following exposure to ADR. ADR resistance may be mediated by a reduction in the level of DNA strand breaks.     

Bronchocentric granulomatosis: a complication of allergic bronchopulmonary aspergillosis.

Hypersensitivity to the fungal antigens of Aspergillus fumigatus may result in a spectrum of immune injury collectively known as allergic bronchopulmonary aspergillosis (ABPA). This report describes a 14-yr-old boy who presented clinical findings consistent with ABPA,including a history of asthma, blood eosinophilia, serum precipitins, and IgE antibodies to Aspergillus fumigatus. Sputum Aspergillus, pulmonary infiltrates, and dual types I and III skin reactions to Aspergillus fumigatus were observed also. Pathology of the resected right upper lobe revealed severe bronchial destruction with the findings of bronchocentric granulomatosis. Noninvasive septate fungal hyphae compatible with Aspergillus were identified. Cultures from sputum and surgical specimens grew Aspergillus and Mycobacterium intracellulare avium. The PPD-B (purified protein derivative-Batty) intradermal skin test produced a 6 mm induration (PPD-S was negative). The patient's condition has been well controlled with prednisone and several antituberculous drugs. In addition, inflammatory and immunologic parameters have begun to return to normal. The relationship between ABa and the atypical mycobacterial infection is not clear. The association of ABPA with the severe bronchial destruction seen in bronchocentric granulomatosis is emphasized to alert physicans to this serious sequelae of ABa seen in the asthmatic.     

Effects of acute and chronic treatment with imipramine on 5-hydroxytryptamine nerve cell groups and on bulbospinal 5-hydroxytryptamine/substance P/thyrotropin releasing hormone immunoreactive neurons in the rat. A morphometric and microdensitometric analysis

Summary Groups of male rats were treated for a period of 14 days with imipramine (10mol/kg) given twice daily. Separate groups of rats received a single dose treatment using the same dose and experimental design as for the repeated treatment. Employing the avidin-biotin immunoperoxidase technique for immunohistochemistry 5-hydroxytryptamine (5-HT)-, substance P (SP)- and thyrotropin releasing hormone (TRH)-like immunoreactivities (IRs) were visualized in consecutive coronal sections of the brain stem and of the spinal cord. The IRs were studied by means of morphometric and microdensitometric procedures using automatic image analysis on profiles representing nerve terminal networks of the ventral horn of the cervical and lumbar enlargements of the spinal cord as well as their coexistence (5-HT/SP and 5-HT/TRH). With the same technique 5-HT IR was measured in the 5-HT nerve cell groups of the medulla oblongata (B 1, B 2, B 3) and of the nucleus raphe dorsalis (B 7) of the midbrain. In addition 5-HT and 5-hydroxyindolacetic acid (5-HIAA) levels were measured in the ventral and dorsal horns of the cervical and lumbar enlargements of the spinal cord using high performance liquid chromatography (HPLC). In the same parts of the spinal cord SP IR was studied by means of radioimmunoassay (RIA).The microdensitometric studies showed that chronic, but not acute, imipramine treatment selectively increased SP IR in the 5-HT/SP/TRH costoring nerve terminals of the medial part of the ventral horn in both the cervical and the lumbar enlargements. Furthermore, quantitative analysis of the entity of coexistence in the 5-HT nerve terminal networks of these areas showed that all the 5-HT nerve terminals contained SP and TRH IRs and that this phenomenon remained after acute and chronic imipramine treatment. The microdensitometric studies on the 5-HT nerve cell groups of the medulla oblongata and of the nucleus raphe dorsalis demonstrated that chronic, but not acute, imipramine treatment selectively increased 5-HT IR in the nerve cell bodies of the lateral part of group B 3 as evaluated from the median grey values. Acute, but not chronic, imipramine treatment significantly increased the field area of 5-HT IR of nerve cell bodies in group B 7, reflecting an increase in the mean profile area of the 5-HT IR nerve cell body profiles. Instead, the mean profile area of 5-HT IR cell bodies of group B 1 was acutely reduced by imipramine.The biochemical studies demonstrated that chronic imipramine treatment selectively reduced 5-HT utilization in the ventral horn of the spinal cord and selectively increased SP IR in the dorsal horn of the lumbar enlargement.In view of these observations it is suggested that chronic imipramine treatment specifically increases SP IR in the 5-HT/SP/TRH costoring nerve terminals of the ventral horn probably related to reduced SP release and reduced 5-HT utilization in these terminals. The results obtained in group B 7 may be explained by a regulation by the³H-imipramine raphe binding sites of fast axonal transport, an influence which may have therapeutic consequences. This mechanism may also be responsible for the increase in 5-HT IR seen upon chronic imipramine treatment in the lateral part of the 5-HT nerve cell body group B 3. Such an effect may lead to a metabolic down-regulation of group B 7, having a possible role for the antidepressant activity of imipramine. The reduction of the mean profile area of 5-HT IR cell bodies of group B 1 seen in the acute treatment can possibly be caused by, noradrenaline (NA) uptake inhibition in inhibitory NA terminals innervating the B 1 group. These results also illustrate the heterogeneities in the responses of the 5-HT nerve cell groups to antidepressant treatment. The ability of chronic imipramine treatment to increase SP IR in the dorsal horn of the lumbar enlargement may reflect the existence of a monoamine-SP interaction in the substantia gelatinosa due to the NA and/or 5-HT uptake blocking activity of imipramine. The existence of such an interaction may help to explain the antinociceptive effect of chronic imipramine treatment.Part of the paper was presented at the 17th International Congress of the International Society of Psychoneuroendocrinology, Bergen, June 29–July 4, 1986.     

Substance P and calcitonin gene-related peptide: immunohistochemical localisation and microvascular effects in rabbit skeletal muscle

Summary 1. The distribution and microvascular effects of substance P (SP) and calcitonin gene-related peptide (CGRP) were studied in the rabbit tenuissimus muscle using immunohistochemistry and intravital microscopy. 2. Individual fibers within nerve bundles and along blood vessels in the muscle were found to be immunoreactive (IR) for both SP and CGRP, thus showing an apparently complete coexistence for these peptides. In dorsal root ganglia most SP-positive cells were also CGRP-IR, but the latter cells were somewhat more numerous than SP-IR cells. 3. When applied topically to the muscle, both SP and CGRP increased blood flow in a dose-dependent manner, but CGRP was more potent and caused responses of longer duration. Both SP and CGRP dilated transverse arterioles, but they had little or no effect on the smaller terminal arterioles. This resulted in a redistribution of blood flow to the connective tissue adjacent to the muscle. 4. SP, but not CGRP, elicited vigorous vasomotion in larger arterioles and caused the formation of aggregates of platelets and leukocytes in the venules. Neither flow increase, nor vasomotion or aggregate formation were influenced by pretreatment of the animals with mepyramine, cimetidine or indomethacin. Capsaicin (1 M) had a powerful effect on transverse arterioles resembling that of both SP and CGRP. 5. It is concluded that some of the vascular effects hitherto ascribed to SP on the basis of nerve stimulation and application of capsaicin might, at least in part, be due to release of CGRP. Send offprint requests to: A. Ohlen, Department of Physiology, Karolinska Institutet, S-104 01 Stockholm, Sweden     

Expression of P53 during lens epithelial cell apoptosis induced by ultraviolet

Cataract is a disease induced by multi- factorsand its pathologic mechanism isstill unknown.Ithasbeen accepted that ultraviolet irradiation ( UVR) isone of the most important pathogenesis.Li etal hasreported that UVR could induce the apoptosis of lensepithelial cells( LECs) and resulted in lens opacities.In order to study the pathologic mechanism of UVR-induced cataract at molecular and cellular levels,weinvestigated the apoptosisof LECs and the expressionof P5 3.1　 MATERIALSAND M…     

Assessment of high-energy phosphorus compounds in the rat kidney by in situ31P nuclear magnetic resonance spectroscopy: effect of ischemia and furosemide

Summary ³¹P nuclear magnetic resonance (NMR) spectroscopy of the in situ rat kidney was performed by a surface coil method, and the effects of ischemia and furosemide infusion were assessed.³¹P NMR spectra of the kidney subjected to 30 min of ischemia returned completely to the pre-ischemic level after 60 min of reperfusion. But the³¹P NMR spectra after 60 min of ischemia did not recover, even after 120 min of reperfusion. Levels of -ATP and inorganic phosphate (Pi) decreased and the chemical shift of Pi increased after intravenous infusion of furosemide. This increase in chemical shift might signal an alkalotic change in intracellular pH. Furosemide infusion prior to ischemia is thought to protect the kidney from injury induced by 60 min of warm ischemia. The chemical shift of Pi returned to the pre-ischemic level earlier than -ATP and Pi. In conclusion, according to the findings of³¹P NMR spectroscopy, furosemide infusion prior to ischemia may be effective in protecting the kidney against ischemic injury. But the change in Pi peak and the causes of the dissociation of Pi and -ATP should be examined further.