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Effects of internal free calcium upon the sodium and calcium channels in the tunicate egg analysed by the internal perfusion technique.

1. The unfertilized egg of the tunicate, Halocynthia roretzi, was intracellularly perfused with various solutions. 2. The perfusion apparatus consisted of lower and upper compartments which were connected by a small glass funnel. A denuded egg cell without chorion was dropped into the funnel and brought into close contact with the glass wall of the funnel. The membrane of the egg faced to the lower compartment was ruptured by a slight difference of hydrostatic pressure and the inside of the egg was perfused with the internal solution flowing through the lower compartment. The current across the upper membrane was analysed by voltage-clamp technique. 3. The egg cell in contact with 400 mM-Na external solution and perfused intracellularly with 400 mM-Na for 30 min showed a relatively low Na reversal potential, +6 mV, in comparison with +60 mV in the intact egg in standard artificial sea water. The exchange efficiency was monitored by observing the shift of Na reversal potential during perfusion with high Na internal perfusate. 4. The internal perfusate containing F- ions stabilized the egg membrane and kept the excitability for 1--2 hr during the intracellular perfusion. With the internal F- perfusate the intracellular cationic content was changed to 400 mM-Na, K, Rb or Cs (external solution of 400 mM-Na) and permeability ratios of the egg Na channel were estimated as PNa:PK:PRb:PCs=1.0:0.14:0.05:0.04. The internal F- perfusate abolished Ca current which was consistently observed in the intact egg, while the internal Cl- perfusate kept both Na and Ca current as in the intact egg. However with the internal Cl- perfusate the egg cell could not be kept in good condition more than 20-30 min. 6. The effects of intracellular free Ca ions upon the egg Na and Ca channels were analysed by using Ca ion-buffered internal Cl- and high Na perfusate. The results showed that internal Ca ions above 10(-6) reduced the Ca current and enhanced the Na current at the same time. In the range between 10(-5) and 10(-4) M the Ca current became half of the control obtained with zero free Ca perfusate while the Na conductance at the zero current level doubled. The internal Ca ions above one mM seemed to abolish the Ca current and to reduce the Na current as well. The reciprocal effect of intracellular Ca ions upon the egg Na and Ca channels was demonstrated in the concentration range from 10(-6) to 10(-3) M.

Animals

Receptor-mediated internalization of bradykinin. DDT1 MF-2 smooth muscle cells process internalized bradykinin via multiple degradative pathways.

This study was undertaken to evaluate the role of internalization in the action of the peptide autacoid bradykinin (BK). At 4 degrees C [3H]BK binds to an apparently single class of B2 kinin receptors on DDT1 MF-2 smooth muscle cells (C. M. Munoz, S. Cotecchia, and L. M. F. Leeb-Lundberg, manuscript submitted). At this temperature the [3H]BK binding was confined exclusively to the cell surface. On the other hand, at 37 degrees C the B2 receptor-specific cell surface [3H]BK binding was rapidly followed by a receptor-specific internalization of [3H]BK (t1/2 approximately 9 min). The internalization reached a steady-state level after 30-40 min that was 80-100% of the level of specifically bound [3H]BK on the cell surface at 4 degrees C, and this level was maintained for greater than or equal to 2 h. Internalized [3H]BK was routed via at least two intracellular degradative pathways which were distinguished primarily based on subcellular localization but also on a small but significant difference in the rate of [3H]BK degradation. One pathway was localized in a plasma membrane-enriched fraction and had a relatively high degradative capacity. Another pathway was localized in a microsomal fraction and had a relatively low degradative capacity. The internalized [3H]BK activity was rapidly released into the media (t1/2 approximately 24 min). Following a single round of internalization, the released activity consisted almost exclusively of small [3H]BK fragments (less than [3H]BK(1-5)). In contrast, at steady-state [3H]BK represented 30-40% of the released activity. While chloroquine (100 microM) did not alter the rate of [3H]BK internalization or release or the intracellular distribution of [3H]BK, this agent significantly decreased the rate of [3H]BK degradation in both pathways. In all, these results show that B2 kinin receptor-mediated internalization of BK is a process integral to the interaction of BK with DDT1 MF-2 smooth muscle cells and may be a mechanism for terminating BK actions by rapidly removing extracellular free and receptor-bound BK and accessing various intracellular BK degradative pathways.

Amino Acid Sequence

Rabies vaccine standardization: International Collaborative Study for the Characterization of the fifth International Standard for Rabies Vaccine.

A collaborative study was carried out to establish a replacement for the International Standard for Rabies Vaccine, the stocks of which are exhausted. Three rabies vaccines for human use derived from different rabies virus strains and prepared on different cell culture substrates were compared with the International Standard for Rabies Vaccine using in vivo and in vitro assay methods in a collaborative study involving 14 participants. The proposed fifth International Standard (PISRAV) which was derived from the same virus strain as the present international standard preparation, the Pitman Moore (PM) strain, was found to be approximately twice as potent relative to the International Standard in immunogenicity assays as in antigenicity assays. On the other hand another vaccine, derived from the LEP strain, was considerably more potent in antigenicity assays than in immunogenicity assays. The glycoprotein of the proposed replacement standard measured in antigenicity assays appeared to be stable at +37 degrees C for 245 days, whereas the immunogenicity of the proposed replacement vaccine was sensitive to this heat treatment and the vaccine lost 66% of its immunogenic potency. The results of this study indicate that the NIH protection test should continue to form the primary basis for potency assay of rabies vaccine as glycoprotein content does not appear to correlate with immunogenic potency for different types of vaccine. The vaccine coded PISRAV has been established as the fifth International Standard for Rabies Vaccine and a potency of 16 International Units of Rabies Vaccine (based on the immunogenicity assays) assigned to the contents of each ampoule. Each ampoule has also been assigned a unitage of 10 IU of PM Rabies Virus Glycoprotein and 135 IU of PM Rabies Virus Ribonucleoprotein.

Animals

Effects of internal potassium and sodium on the anomalous rectification of the starfish egg as examined by internal perfusion.

1. The effects of alterations of the intracellular ionic composition on the properties of anomalous (or inward) rectification of the egg membrane of the starfish, Mediaster aequalis, were studied by using an intracellular perfusion technique. The following results were obtained, analysing the membrane current with the voltage-clamp technique. 2. The inward rectification of the K conductance depends only on the membrane potential, V, when the K equilibrium potential, VK, is altered by changing the internal K+ concentration at a fixed external K+ concentration, while it depends on V-VK when VK is altered by changing the external K+ at a fixed internal K+ concentration. 3. From the above the conclusion is reached that the gating of the K channel of the inward rectification depends on V and external but not internal K+ concentration. 4. The conductance of the K channel at a given voltage is roughly proportional to the square root of [K+]i when the latter is altered at a fixed external K+ concentration. 5. Since the conductance is proportional to the square root of [K+]o when this is altered at a fixed internal K+ concentration, the final conclusion is that this conductance is proportional to the geometric mean of the external and internal K+ concentrations. 6. Intracellular Na+ ions are necessary for the activation of inward rectification; the K conductance increases sharply with internal Na+ concentration, reaching saturation at 200 mM. 7. A similar potentiating effect is found for Li+, although it is weaker. Rb+, Cs+ and organic cations such as arginine+ do not have this effect.

Animals

Internalization of glycosyl-phosphatidylinositol (GPI)-anchored lymphocyte proteins. II. GPI-anchored and transmembrane molecules internalize through distinct pathways.

Ly-6A.2 (T cell-activating protein, TAP) and Thy-1 are glycosyl-phosphatidyl-inositol (GPI)-anchored proteins expressed on the surface of murine T lymphocytes. We have found that Ly-6A.2 (TAP) and Thy-1 are internalized by T cells. In the present study we have investigated whether these GPI-anchored proteins enter cells by endocytosis through coated pits. Two lines of evidence argue against the involvement of coated pits in the internalization of Ly-6A.2 (TAP) and Thy-1. First, drugs that effectively blocked the endocytosis of transferrin receptor and H-2 class I molecules, (which are known to be internalized via coated pits) did not inhibit the internalization of the GPI-anchored proteins. Second, in ultrastructural analyses, Ly-6A2 (TAP) and Thy-1, in contrast to the transferrin receptor, were rarely found in coated pits or vesicles. These observations suggest that the GPI-anchored proteins on T lymphocytes are internalized by a distinct pathway that does not involve endocytosis through coated pits.

Animals

Internal medicine and the training of international medical graduates: a time for open discussion and new approaches.

The number of foreign national medical graduates entering internal medicine residency training programs in the United States has doubled since 1986. A rigorous, standardized preresidency evaluation of the basic clinical skills and language abilities of international medical graduates should be implemented. Those found to have significant deficits should undertake a preparatory curriculum designed to meet special educational needs before entry into the formal training program. A relevant curriculum might include formal lectures, reading assignments, physical diagnosis sessions, language classes, patient encounter sessions, and a tutorial on U.S medical culture that includes medical ethics and the basics of the our health care system. All or only some of these may be required for any given individual. The Educational Commission for Foreign Medical Graduates (ECFMG) could provide many of the methods needed for an evaluation program and work collaboratively with program directors. This new approach to training of international medical graduates will require an evaluation system to to measure its effectiveness. Important questions about the funding of graduate medical education for international medical graduates must also be addressed.

Clinical Competence

[A giant aneurysm of the internal carotid artery treated by intracranial direct surgery--with a special reference to the anatomical relationship between the cavernous sinus and the internal carotid artery (author's transl)].

There have been a few reports about intracranial giant aneurysms treated by intracranial direct approaches. Especially, for giant aneurysms of the internal carotid artery, direct operation has been thought to be difficult to perform because of the anatomical particularity and the danger of rupture during surgery. So the cervical carotid ligation has frequently been indicated. However, the carotid ligation does not relieve the symptoms caused by the giant aneurysm as an "intracranial mass lesion". The authors have reported here, a cured case of a giant aneurysm of the intracavernous portion of the internal carotid artery, to which, as the first step, ligation of the internal carotid artery in the cervical region was performed, and as the second step, endaneurysmorrhaphy was carried out for the remaining symptoms caused by the "intracranial mass lesion". Furthermore, the surgical techniques for treatment of giant aneurysms of the internal carotid artery were discussed citing literatures, and the authors concluded that the combined operation of carotid ligation and endane-urysmorrhaphy could be effective to giant aneurysms in this region. Meanwhile, a new concept concerning the anatomy of the carotid-cavernous region already proposed by Bedford, was confirmed by our observation at operation. It may be considered that the intracranial direct approach to aneurysms in this region and also to carotid cavernous fistulae should more frequently be indicated than ever performed, according to the new concept of the anatomy in this region.

Adult

Studies on phage internal proteins: formation of internal protein - T2 DNA complexes in vivo.

Internal proteins, synthesized in T2-infected Escherichia coli B cells were recovered from bacterial membranes during the early stages of infection. Approx. 15 min after the onset of infection, T2 and T4 internal proteins were released from the bacterial membranes and sedimented along with newly synthesized phage DNA. Internal protein-DNA complexes were also obtained by chromatography on hydroxylapatite columns. Internal proteins were not released from bacterial membranes after infection with amber mutants defective in genes 21, 22 and 23. It has been suggested that these complexes are intermediates in T-even phage morphogenesis.

Cell Membrane

International collaborative study by in vitro bioassays of the first international standard for porcine inhibin.

A lyophilized preparation of inhibin from porcine ovarian follicular fluid, ampoule code 86/690, was made internationally available as a research standard for in vitro bioassays in 1987. A study involving ten participants in eight countries assessed the stability and suitability of this research standard to serve as an international standard. Each of the participants used in vitro assays, the majority of which depended upon the inhibition of release of follicle-stimulating hormone from dispersed rat anterior pituitary cells. The research standard 86/690 was compared with coded ampoules of 86/690 stored under conditions of accelerated thermal degradation and with inhibins from different species. Intra- and interlaboratory variation for estimates of potency of a coded duplicate ampoule of the research standard provided the basis for comparisons of non-identical inhibins, but the fourfold variability of potency estimates for identical ampoules was such that no conclusions about the differences seen for non-identical inhibins could be made. Predictions of stability from consensus estimates of potency of ampoules that have undergone accelerated thermal degradation indicated that the research standard had satisfactory stability. On the basis of this study, the research standard 86/690 was deemed sufficiently stable and suitable to serve as a standard for in vitro bioassays and was established by the World Health Organization Expert Committee on Biological Standardization as the First International Standard for Porcine Inhibin. The possible presence, in biological extracts (standard or sample), of other bioactive proteins, such as activin and follistatin, complicates the quantitative interpretation of bioassay data. A standard of highly purified human inhibin is now required as a standard for immunoassays used for clinical research purposes; sufficient quantities of recombinant human inhibins have recently been donated for ampouling and evaluation by bio- and immunoassay in the subsequent phase of the standardization of inhibins.

Animals

A national study of internal medicine and its specialties: I. An overview of the practice of internal medicine.

A nationwide study of 24 medical and surgical specialties has been conducted by the University of Southern California School of Medicine, Division of Research in Medical Education. This article is the first in a series reporting findings for general internal medicine and 10 subspecialties of internal medicine. Populations for these 11 specialties are defined and enumerated, and the specialties are compared in terms of demographic and geographic distribution. Practice comparisons are presented based on characteristics such as workload, allocation of professional time, location of encounters with patients, distribution of primary problem diagnoses, and projections of annual patient encounters. Forthcoming specialty-specific articles will present highly detailed information for general internal medicine and for the subspecialties of cardiology, gastroenterology, pulmonary disease, allergy, hematology, endocrinology, nephrology, medical oncology, rheumatology, and infectious diseases.

California

A national study of internal medicine and its specialties: II. Primary care in internal medicine.

A nationwide study of practitioners in 24 medical and surgical specialties was conducted by the University of Southern California School of Medicine, Division of Research in Medical Education. In this second report of a series presenting findings for internal medicine, general internal medicine and 10 subspecialties of internal medicine are compared using a care classification scheme designed for the study. On the basis of characteristics of the individual patient encounter, this care classification scheme distinguishes several dimensions associated with the concept of primary care. Five empirically derived types of care rather than a simple "primary" or "non-primary" dichotomy are described, and the distributions of each type for the 11 subspecialties examined are noted. Types of care (according to the care classification) are examined by the time per patient, the complexity of physician services, the severity and chronicity of problems, and the degree of specialization associated with providing different types of care. Estimates of the number of annual encounters, and the number of annual encounters for three of the five types of care, with the proportion of each accounted for by each subspecialty, are given.

Consultants

[Internal maxillary artery and collateral circulation in thrombosis of internal carotid artery (author's transl)].

From 53 cases of thrombosis of the internal carotid artery in the neck, collateral circulation through external carotid branches was angiographically demonstrated in 34 subjects. Anastomosis between external and intracranial circulations was seen to take place through three ways formed by internal maxillary, superficial temporal and facial arteries. The role of internal maxillary artery is emphasised as the most important pathway in the connections between the two circles.

Adult

Graduated internal dilatation in the treatment of fibromuscular dysplasia of the internal carotid artery.

A patient with transient ischaemic attacks due to fibromuscular dysplasia of the internal carotid artery is presented. Standard methods of vascular reconstruction may not be possible as the disease often involves the distal half or two-thirds of the extracranial internal carotid artery. For this reason the technique of graduated internal dilatation first described by Morris et al. was employed and showed satisfactory results on short-term follow-up.

Adult

International standard for anti-Brucella abortus serum: comparison of the complement-fixing activity of the first and second international standards and the EEC standard.

Although the three standards have similar agglutinating activities, it was found that the first International Standard had about half the complement-fixing activity of the second International Standard and the EEC standard. The complement-fixing activities of the latter two standards were similar. On the basis of these results, showing that the agglutination and complement-fixation tests measure different biological activities in anti-Brucella abortus sera, the WHO Expert Committee on Biological Standardization in 1976 assigned separate unitages of agglutinating and complement-fixing activity to the second International Standard for Anti-Brucella abortus Serum.

Agglutination Tests

An International Collaborative Study of the assay of human placental lactogen: Establishment of WHO International Reference Preparation of human placental lactogen.

A reference preparation of human placental lactogen (HPL) for use in immunoassays is described. In an international collaborative study is was found to be stable and suitable for use as a standard for HPL in plasma. Estimates of HPL in plasma made in different laboratories agreed more closely when expressed in terms of this reference preparation than in terms of individual laboratory standards. The reference preparation is available in ampoules (coded 73/545) which each contain an identical amount of HPL, found in the study to be approximately 850 microgram. The preparation has been established as the International Reference Preparation for HPL for immunoassay and, based on the results of the study, the content of each ampoule is defined as 850 micro International Units.

Female