Spontaneous and agonist-induced openings of an acetylcholine receptor channel composed of bovine muscle α-, β and δ-subunits

During the development of mammalian muscle the -subunit of the nicotinic acetylcholine receptor (AChR) is replaced by the -subunit to produce well-defined alterations in the conductance and gating of the channel. To gain a better unterstanding of the functional role of the and -subunits, we have studied the properties of an AChR channel lacking these subunits. The AChR expressed in Xenopus oocytes injected with the bovine -, and -subunit-specific mRNAs (referred to as -AChR) is unusual in that its channel opens spontaneously at a high frequency in the absence of agonist. From a comparison of the -AChR with complete receptors containing either the or -subunit, we conclude that the and -subunits influence most channel properties, including agonist binding, and are especially important for stabilizing the closed state of the unliganded receptor channel. The -AChR can form when a complete set of four subunit-specific mRNAs is injected. The ease with which it is assembled raises the possibility that the - AChR contributes to some of the variations in receptor properties that occur during development.     

Recognition by peptide mapping of three different structural groups of outer membrane protein YOP-1 of Yersinia enterocolitica and Yersinia pseudotuberculosis

The structural relation of YOP-1 of european and american Yersinia enterocolitica serotypes O3, O9, O5, 27, and O8 and O20, respectively, and Y. pseudotuberculosis serotypes I, II, and III was compared by sodium dodecyl sulfate polyacrylamide gel electrophoresis and peptide mapping using Staphylococcus aureus protease V8. Apparent molecular weights of YOP-1 ranged from 206,000 (O3) to approx. 180,000 (O8). According to their respective peptide maps YOP-1 of the european and american Y. enterocolitica serotypes and Y. pseudotuberculosis serotypes could be assigned to three different groups. Evaluation of several isolates of Y. enterocolitica serotypes O3, O9, and O8 by peptide mapping indicated that YOP-1 is conserved within a serotype. However, one serotype O8 isolate differed from the consensus peptide pattern of the other serotype O8 and O20 isolates. The similarity of the peptide patterns of Yersinia serotypes which predominate in certain geographical locations, i. e., european and american Y. enterocolitica serotypes, suggest common evolution of YOP-1 of these serotypes independent of the evolution of the other serotypes.     

Analysis of cytosolic and lysosomal pH in apoptotic cells by flow cytometry

Several reports indicate that the cytosol is acidified during apoptosis although the mechanism is not yet fully elucidated. The most acidic organelle found in the cell is the lysosome, raising the possibility that lysosomal proton release may contribute to the cytosolic acidification. We here describe methods for measurement of the cytosolic and lysosomal pH in U937 cells by a dual-emission ratiometric technique suitable for flow cytometry. Cytosolic pH was analysed in cells loaded with the fluorescent probe BCECF, while lysosomal pH was determined after endocytosis of FITC-dextran. Standard curves were obtained by incubating cells in buffers with different pH in the presence of the proton ionophore nigericin. Apoptosis was induced by exposure of cells to 10ng/ml TNF- for 4h, and apoptotic cells were identified using a fluorescent marker for active caspases. By gating of control and apoptotic cells, the cytosolic and lysosomal pH were calculated in each population. The cytosolic pH was found to decrease from 7.2 ± 0.1 to 5.8s±0.1 and the lysosomal increased from 4.3±0.4 to 5.2±0.3. These methods will be useful in future attempts to evaluate the involvement of lysosomes in the acidification of the cytosol during apoptosis.     

Synergistic effects of tumour necrosis factor-α and interferon-γ on feline haemopoietic progenitors

Pathogenic mechanisms that underlie feline leukaemia virus subgroup-C (FeLV-C) induced erythroid aplasia are unknown. FeLV-C infection is associated with higher serum levels of interferon- (IFN-) and tumour necrosis factor- (TNF-), which may act synergistically to cause haemopoietic suppression. In the present studies, the synergistic effects of TNF- and IFN- on feline bone marrow progenitors in vitro were evaluated. Bone marrow mononuclear cells from specific-pathogen-free cats were exposed to TNF- (100 and 200 pg/ml) and IFN- (100 or 200 units/ml), alone or in combination, for 2 h before plating for clonal assays of colony forming units. Our results show that TNF- and IFN- in combination caused marked suppression of feline colony forming units-erythroid (CFU-E), burst forming units-erythroid (BFU-E), and colony forming units-fibroblasts (CFU-F), whereas colony forming units-granulocyte/macrophage (CFU-GM) were minimally affected. The same concentrations of TNF- and IFN- alone had minimal effects on CFU-E, BFU-E and CFU-F. These results suggest that TNF- and IFN- may play a significant role in regulating haemopoiesis in cats and may be involved in the pathogenesis of erythroid aplasia in cats infected with feline leukaemia virus.     

Evidence for a direct relationship between mitochondrial genome organization and regeneration ability in hexaploid wheat somatic tissue cultures

Summary Embryogenic and non-embryogenic long-term callus cultures of hexaploid wheat exhibit differences in the organization of their mitochondrial genome. Embryogenic and non-embryogenic fractions of callus cultures initiated from immature embryos of the wheat cultivar Chinese Spring have been isolated and subsequently subcultured. DNA-DNA hybridization experiments using labelled cloned wheat mitochondrial DNA fragments have shown that the mitochondrial DNA organization of embryogenic subcultures derived from embryogenic parts of Chinese Spring calli is closely related to that of the initial Chinese Spring calli, while non-embryogenic subcultures derived from non-embryogenic fragments of Chinese Spring calli exhibit a mitochondrial DNA organization similar to that found in non-embryogenic calli derived from cultivar Aquila. In addition, somatic tissue cultures initiated from three other non-embryogenic wheat cultivars (Talent, Thésée and Capitole) display mitochondrial DNA arrangements similar to those found in cultivar Aquila. These results strongly suggest that, in wheat callus cultures, a particular mitochondrial genome organization is correlated with the ability of cultured cells to regenerate whole plants.Abbreviations mtDNA mitochondrial DNA - ctDNA chloroplast DNA - rRNA ribosomal RNA - kb kilobase pair - cv cultivar - 2,4-D 2,4-dichlorophenoxyacetic acid     

Expression of growth factor receptors in injured nervous tissue. I. Axotomy leads to a shift in the cellular distribution of specific beta-nerve growth factor binding in the injured and regenerating PNS

Summary We have studied -nerve growth factor (-NGF) receptor expression in the injured and regenerating chick PNS using [¹²⁵I]-iodinated -NGF as a radioactive probe to map and quantitate autoradiographically thein situ distribution of specific [¹²⁵I] -NGF binding.Two different mechanisms are involved in the reappearance of specific [¹²⁵I] -NGF binding on the normally unlabelled adult peripheral nerves. The anterograde and retrograde axonal transport of -NGF binding sites leads to a rapid but transient accumulation of [¹²⁵I] -NGF binding on both sides of crushed or transected sciatic and brachial nerves. There is a dramatic decrease in the axonal transport of -NGF binding sites, starting 1 day after, nerve injury (1 DPO) and reaching basal levels of 10–20% of the control values at 3 to 10 DPO. Gradual but complete recovery of this axonal transport was noted in the sciatic neurites allowed to regain contact with their peripheral targets. A very different regulation pattern was observed for the local reappearance of specific [¹²⁵I] -NGF binding on the endoneurial Schwann cells throughout the distal part of the axotomized nerve. It was first observed at 4 DPO, becoming maximal at 6 DPO. Reinnervation of the nerve after crush led to a rapid decrease of this specific [¹²⁵I] -NGF binding, which followed a proximo-distal temporal gradient.These results show that axotomy leads to a drastic decrease in the axonal expression of [¹²⁵I] -NGF binding, while causing its appearance on the Schwann cells of the denervated endoneurium. They suggest that these endoneurial cells may become the primary target for -NGF following axotomy and during regeneration.     

Übererregbarkeit der γ-Motoneurone beim „Stiff-man“-Syndrom

Zusammenfassung Es wurde eine 40jährige Frau mit sog. Stiff-man-Syndrom untersucht. In Narkose und Lumbalanaesthesie verschwand die abnorme Muskelverspannung. Ebenso normalisierte sich unter Differentialblockade des N. femoralis durch Procainumspritzung der Tonus im Quadriceps vorübergehend ohne Lähmungserscheinungen. Daraus kann gefolgert werden, daß die-Motoneurone wesentlich am Zustandekommen der Muskelverspannung beteiligt sind. Histologisch ergab sich kein krankhafter Muskelbefund. Elektromyographisch bestand eine langanhaltende Aktivität in den befallenen Muskeln, die sich von Willkürentladungen nicht unterscheiden ließ. Bei passiven Bewegungen wurde eine ungewöhnliche Entdehnungsaktivität registriert. Unter Willkürinnervation bestand ein normal dichtes Muster. Bei Entspannung nach aktiver Innervation trat nach kurzer Entlastungsreaktion Überdauerungsaktivität auf. Einzelpotentiale, Nervenleitgeschwindigkeit, mechanisch und elektrisch (H-Reflex) ausgelöste Muskeleigenreflexe waren normal, die silent period nicht verändert. Der H-Reflex zeigte unter subparalytischer Dosis von Succinylcholin einen Abfall wie beim Gesunden.Unter Behandlung mit Diazepam und einem Amino-Buttersäurederivat verschwand die abnorme Muskelspannung fast vollständig. Es wird die Annahme vertreten, daß es sich bei dieser Form des Stiff-man-Syndroms, die dem Syndrom vonMoersch undWoltman entspricht, um ein Parabiosephänomen der-Motoneurone handelt (Spindelmyotonie). Analog dazu liegt beim Isaacs-Syndrom eine Überaktivität der peripheren -Motoneurone vor (Neuromyotonie). Bei anderen in der Literatur beschriebenen Formen des Stiff-man-Syndroms handelt es sich offensichtlich um primäre Muskel- oder Bindegewebserkrankungen.

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Summary A 40 years old woman suffering from the so-called stiff-man syndrome is described. The abnormal muscle-tonus disappeared in narcosis and in lumbar anaesthesia. When procaine was infiltrated close to the nervus femoralis, the tonus in the quadriceps muscle was normalised temporarily without the muscle being paralysed. Consequently the-motoneurones may be considered as an essential factor in bringing about the pathological muscle stiffening. Histologycally there was no evidence of muscle disease. The EMG showed a long-lasting activity in the stiffened muscles which did not differ from a normal pattern. During passive shortening of the muscle abnormal activity was registered. An interference pattern was seen with maximum voluntary contraction. Active innervation was followed by anew electrical activity in the relaxing muscle, only interrupted by a short silent interval. Single units, nerve conduction velocity, the potentials of the muscle stretch reflex an the H-reflex were normal. The silent period was not altered. Under subparalytical dosage of succinylcholine the amplitude of the H-reflex showed a decrease in a physiological manner. The muscle stiffness disappeared almost completely when the patient was treated with diazepam (Valium) and a derivate of amino-butyric acid (CIBA 34-647 Ba).There is some reason to believe that this variety of stiff-man syndrome, being identical with the syndrome described byMoersch andWoltman, is caused by the parabiotically diseased-motoneurones (spindlemyotonia). Likewise a hyperactivity of peripheral -motoneurones is known to bring aboutIsaacs syndrome (neuromyotonia). Other varieties of the stiff-man syndrome described by literature are obviously primary diseases of the muscle or connective tissue.

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Herrn ProfessorG. Schaltenbrand zum 70. Geburtstag gewidmet.     

An improved apparatus for the optical recording of contraction of single heart cells

An optical system for measuring changes in cell length during unloaded contractions of cardiac myocytes is described. A one-dimensional video image of a cell is obtained every 4 ms with a linear photodiode array, which is aligned with the longitudinal axis of the cell. The circuit used to process the image from the photodiode array has a variety of features to aid in the accurate determination of the distance between the ends of the cell, i.e. the cell length. First, the video image of the cell is divided into two windows, one encompassing the front edge of the cell, the other encompassing the rear edge. Other cells or debris beyond the cell edges are excluded. Changes in the general light level, for example as a result of debris floating above the cell, have little effect because within the windows the background light level is subtracted from the signals before they are processed further. To detect the cell edges, the system determines when the signals within the windows exceed (front edge) or drop below (rear edge) chosen threscholds, which are different for the front and rear edges. The system has memory and it identifies the rear edge of the cell as the last time the signal falls below the threshold; because of this bright spots within the cell are not mistaken for the end of the cell. The system has hysteresis, which enables it to ignore small fluctuations in brightness around the threshold. The system is easy to use, accurate, readily calibrated, and it has good spatial and time resolution (about 0.25 m and 4 ms respectively).     

Convergent inputs from articular,cutaneous and muscle receptors onto ascending tract cells in the cat spinal cord

Summary 1.Responses were recorded from 160 ascending tract cells in segments L4 to L6 of the spinal cord in chloralose anaesthetized, spinalized cats. The tract cells were identified by antidromic activation following stimulation of pathways in the lateral and ventral funiculi at the level of the spinal cord transection at the thoracolumbar junction. Axonal conduction velocities ranged from 9 to 114 m/s. 2. A sample of 152 of the neurones examined could be subdivided according to the distribution of their receptive fields into 49 cells activated just from receptors located in skin (s cells), 17 neurones excited by receptors in deep tissues (d cells), 15 units with a convergent input from receptors in skin and deep tissues (sd cells), and 25 neurones with a convergent input from the knee joint and either skin (sj cells), deep tissues (dj cells) or both (sdj cells). No receptive fields could be demonstrated for the remaining 46 neurones. 3. S and sj cells were found almost exclusively in the dorsal horn, whereas many d, sd, sdj and dj units were in the ventral horn. Almost all of the cells that lacked receptive fields were in the ventral horn or intermediate grey. 4. Ninety-one of 158 cells (56%) demonstrated no background activity. Of these, 43 cells (27%) lacked receptive fields. Many of the silent neurones were in the ventral horn, but some were in the dorsal horn. Of 25 cells having knee joint input, 18 (72%) had background activity. 5. All of the neurones that had a receptive field in the knee joint also had a convergent input from receptors in other tissues. In 3 cases, there was a receptive field in the skin over the foot (sj cells). For 16 cells, receptive fields included not only the knee joint but also skin and deep tissue (sdj cells). Usually, the cutaneous receptive field was near the knee joint, but sometimes it was remote, such as on the foot. The deep receptive fields were chiefly in the muscles of the thigh and/or leg. For 6 dj cells, the receptive fields included not only the knee joint but also deep fields like those of sdj cells. 6. Cutaneous receptive fields were classified as low threshold (cells excited best by innocuous intensities of mechanical stimulation), wide dynamic range (cells activated by weak mechanical stimuli, but the best responses were to noxious stimuli) or high threshold (innocuous stimuli had little effect, but noxious mechanical stimuli produced a vigorous discharge). Similarly, stimulation of the knee joint with weak mechanical stimuli could excite some neurones, while others could be activated by weak or strong articular stimuli but were excited best by noxious stimuli, and still other neurones were activated by knee joint stimuli only if the intensity was noxious. 7. In several instances, contralateral receptive fields were noted. These were generally in deep tissue or in the knee joint. 8. It was concluded that many of the responses to articular stimulation of the spinal cord ascending tract cells examined in this study could have been mediated by the fine afferent fibres that supply the knee joint. Although further work will be required to determine which particular ascending tracts transmit nociceptive information concerning the knee joint, it can be proposed that many of the responses demonstrated here were likely to play a role in either joint pain of in triggering responses associated with joint pain.     

Analysis of the effect of barium ions on isolated medullated nerve fibers

Zusammenfassung An der isolierten markhaltigen Nervenfaser wird nach Ba⁺⁺-Zusatz eine Verschiebung der Elektrotonuskurve nach rechts und unten sowie eine Erhöhung des durch einen Einwärtsstrom bestimmter Größe bewirkten Elektrotonuspotentiales gemessen.Diese Ergebnisse werden im Zusammenhang mit früheren Messungen so gedeutet, daß Barium eine sofortige und anhaltende, der Calciumwirkung vergleichbare Erhöhung der Schwelle bewirkt und daß sich dieser Wirkung eine langsam zunehmende Schwellenerniedrigung überlagert, die durch die Abnahme der Kalium permeabilität verursacht wird.Mit 4 Textabbildungen.     

Modelling of electrolyte transport in renal and intestinal epithelia

Summary Epithelia can be classified as leaky and tight epithelia due to their conductive properties and their modes of solute transport. Both the proximal segment of the nephron and the intestinal tract are leaky whereas the distal nephron and the colon are tight. Consequently, inborn errors and exogenous disorders of solute transport often involve both the proximal tubule and the small intestine. In addition, effects on ion and water transport in the distal nephron closely resemble those in the large intestine. Models of solute transport in leaky and tight epithelia are presented employing porter systems known in mammalian tissues. These porter systems are discussed as possible sites of transport defects and as targets for pharmacological agents.     

Further observations on the morphogenesis of human rotavirus in MA 104 cells

Summary Human rotavirus KUN strain was cultivated in a fetal rhesus monkey kidney cell line, MA 104 cells. Four types of virus particles in cells infected with KUN strain were clearly identified: nucleoid cores, single-shelled particles, double-shelled particles, and membrane band, enveloped particles. Enveloped particles were found only in the thin sections of infected cells. When first visible, the virus precursors appeared at the ribosome free membrane of rough endoplasmic reticulum (RER), increasing in size while simultaneously being coated with nucleocapsid, inner shell. Single-shelled particles were also synthesized within bundles of filaments of viroplasm in the cytoplasma. During subsequent virus maturation two types of budding processes were observed. Double-shelled particles arising at the RER membrane entered the cisternae of the RER through an exocytosis-like process. In contrast, the enveloped particles developed in the cisternae by being completely enclosed with RER membrane, and later during cytolysis released the single-shelled particles. These enveloped virus particles appeared to be the result of inefficient virus maturation at the last stage of outer capsid formation.With 11 Figures     

Cortico-cortical connections of two electrophysiologically identified arm representations in the mesial agranular frontal cortex

Summary Neuronal tracers (diamidino yellow or wheat germ agglutinin conjugated with horseradish peroxidase) were injected in the arm representations of area 6a (mesial surface, area F3), in the arm representation of area 6a (mesial surface) as well as in the eye field of area 6a (dorso-medial surface). The results showed that the arm representation of area F3 receives topographically organized afferents from motor and premotor areas (areas F1, F2, F4 and F5). A further connection was found with that part of cingulate cortex that sends projections to the spinal cord. In contrast, the arm representation of area 6a receives afferents chiefly from area F5, the prefrontal cortex and that part of cingulate sulcus which has few, if any, connections with the spinal cord. No connections were found with the precentral motor cortex (area F1). The area 6a eye field receives afferents mostly from the frontal eye field. Further connections are with the prefrontal cortex and cingulate gyrus. It is suggested that the so called low level motor functions of supplementary motor area are due to the activity of area F3, whereas the so called high level motor functions depend upon an independent area located in area 6a.     

Transfer of tumor specific immunity with “immune” RNA: prospects for the treatment of human cancer

Summary Ribonucleic acids extracted from specifically sensitized lymphoid cells (I-RNA) have been shown to transfer specific immunoreactivity to normal non-immune lymphoid cells. Evidence for the transfer by I-RNA, of immune responses to tumor-associated antigens of animal and human neoplasms, in vivo and in vitro, is reviewed. Results obtained in our laboratory and in other laboratories indicate that xenogeneic, allogeneic and syngeneic I-RNA extracts mediate specific cytotoxicity to tumor cells, in vitro, and mediate transplantation resistance and tumor rejection responses in vivo. Our results suggest that I-RNA preparations fail to elicit immune responses directed against self antigens. By contrast, I-RNA's directed against non-self tumor-associated antigens appear to induce lymphocytes to effect specific anti-tumor immune responses. The mechanisms responsible for the failure of I-RNA to initiate immune responses against self antigens are not known at present and demand investigation.Preliminary results of a clinical Phase I trial of immunotherapy with xenogeneic I-RNA in selected cancer patients are reviewed. I-RNA might offer promise as a new modality for the immunotherapy of human cancer.Recipient of a postdoctoral fellowship from the Deutsche Forschungsgemeinschaft     

Facilitation by 5-hydroxytryptamine of ATP-activated current in rat pheochromocytoma cells

The effects of 5-hydroxytryptamine (5-HT) on an inward current activated by extracellular ATP were investigated in rat pheochromocytoma PC12 cells. Under whole-cell voltage-clamp conditions 5-HT (10 M) reversibly enhanced the amplitude of the current activated by 30 M ATP. The enhancement may not be due to an increase in the number of functional channels because the current activated by 300 M ATP was not remarkably augmented compared with the current activated by 30 M ATP. The current enhancement by 100 M 5-HT was less obvious than that by 10 M 5-HT. When the current kinetics were compared, activation of the ATP-evoked current was accelerated to the same extent by either 10 or 100 M 5-HT, whereas deactivation was largely more accelerated by 100 M 5-HT. Propranolol (10 M), a 5-HT₁ receptor antagonist, or LY53857 (10 M), a 5-HT₂ receptor antagonist, exerted an agonistic effect: the ATP-activated current was facilitated by these compounds. Metoclopramide (10 M), a 5-HT₃ receptor antagonist, neither facilitated the ATP-activated current, nor blocked the current facilitation by 5-HT. Guanosine 5-O-(2-thiodiphosphate) (GDP[S]) (2 mM), the non-hydrolysable analog of guanosine 5-triphosphate (GTP), or K-252a (2 M), a protein kinase inhibitor, did not affect the facilitation by 5-HT of the ATP-activated current when they were included in the intracellular solution. The ATP-activated current pre-facilitated by 10 M dopamine was not enhanced by 10 M 5-HT. Similarly, the pre-facilitation by 5-HT attenuated the current enhancement by dopamine. The results suggest that 5-HT facilitates the ATP-activated channels through receptors that are not readily classified into conventional subclasses of 5-HT receptors. The reciprocal masking between the current facilitation by 5-HT and that by dopamine, combined with their sensitivities to the compounds involved in the intracellular solution, indicates that the facilitation by 5-HT may share not all, but some, common cellular mechanism with that by dopamine.     

Comparison of abilities of recombinant interleukin-1α and -β and noninflammatory IL-1β fragment 163–171 to upregulate C3b receptors (CR1) on human neutrophils and to enhance their phagocytic capacity

Both recombinant IL-1 and - caused an upregulation of C3b receptors (CR1) on human neutrophils and caused a receptor-mediated enhancement of phagocytosis of C3b·IgG-coated microspheres by these leukocytes. The and forms of the recombinant cytokine were of comparable potency regarding CR1 upregulation, although both generally had less than 25% of the potency of FMLP in this respect. Recombinant IL-1 was slightly more potent than the form of the cytokine regarding phagocytosis of opsonized microspheres and, again, both forms were less potent than FMLP in causing an enhancement of phagocytosis by neutrophils. The synthetic noninflammatory immunostimulatory nonapeptide corresponding to residues 163–171 of IL-1 was completely inert with respect to upregulation of CR1 on neutrophils and the enhancement of phagocytosis by these cells. Thus this domain in the intact IL-1 molecule apparently is not involved in CR1 upregulation and the ensuing enhancement in phagocytosis by neutrophils, although it is apparently important in the immunostimulatory activity regarding the proliferation of lymphocytes.     

Expression of estrogen receptors-α and -β in primary breast neoplasms and tumors exposed to neoadjuvant hormonal therapy

We studied the content and expression of mRNA for estrogen receptors receptors- and - in breast tumors before and after 3-month neoadjuvant hormone therapy with antiestrogen tamoxifen and/or aromatase inhibitors. Expression of estrogen receptors- and - was most often detected in ER⁺PR⁺ tumors and most significantly decreased in these neoplasms after exemestane therapy. Immunocytochemical and radioligand assays showed that tamoxifen and anastrozole have little effect on the number of estrogen receptors- The number of progesterone receptors in tumors decreased by the end of anastrozole therapy. Estrogen receptors- were immunocytochemically revealed in 50% primary breast tumors. Anastrozole slightly decreased, while tamoxifen increased the incidence of these receptors. Interruption of signaling through estrogen receptors and suppression of estrogen biosynthesis had different effects on the receptor status of neoplasms and distribution of estrogen receptors- and -.Translated from Byulleten Eksperimentalnoi Biologii i Meditsiny, Vol. 138, No. 11, pp. 559–562, November, 2004     

Eine neue Bestimmung der Neutralfette im Blutserum und Gewebe

Zusammenfassung Die Neutralfette im Blutserum und Gewebe lassen sich über Glycerin exakt ermitteln. Freies und nach alkoholischer Verseifung nachweisbares Gesamt-Glycerin werden mit Glycerokinase in einer dreistufigen enzymatischen Nachweisreaktion im Mikroverfahren bestimmt. Die Differenz wird als Glycerid-Glycerin angesprochen. Reaktionsprinzip, Durchführung der Methode und Berechungsweise werden ausführlich besprochen.Für freies Glycerin werden 0,2 ml Serum, für Gesamtglycerin etwa 0,007–0,015 ml Serum äquivalente Hydrolysatmengen in den Test eingesetzt. Bei einer Empfindlichkeit von 0,0008 µMol (wenn bei 340 nm, bzw. 0,0015 µMol Glycerin, wenn bei 366 nm gemessen wird), können noch 1/20 bis 1/10 des im Normalserum vorhandenen freien bzw. Gesamtglycerins mit den Standardansätzen erfaßt werden. Die obere Nachweisgrenze reicht dann bis 0,15 bzw. 0,3 µMol Glycerin, das entspricht der 8- bzw. 15fachen Normalmenge für freies und Gesamtglycerin. In den angegebenen Grenzen liegen die über Glyceridglycerin nachweisbaren Triglyceridkonzentrationen. Empfindlichkeit und obere Nachweisgrenze können durch andere Serum- bzw. Hydrolysatmengen in weiten Grenzen variiert und der jeweiligen Fragestellung angepaßt werden.

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Summary Triglycerides in blood serum and tissue can be identified exactly by means of glyceride-glycerol. Free and total glycerol, measurable after alcoholic saponification, are determined by glycerolkinase in a three-step enzymatic proof-reaction by means of micro-methods. The difference between total and free glycerol is called glycerideglycerol. Reaction principles, working instructions and the calculation will be discussed in detail.The sensitivity of the standard test runs to 0,0008 respeccively 0,0015 µMol glycerol or 1/20 resp. 1/10 of the normal contentrations. The upper proof limit is at 0,15 respectively 0,3 µMol glycerol per test. This is equal to 8–15 times more than the normal of free and total glycerol. Sensitivity and upper proof limit can easily by varied and adapted to the formulation of the question.

     

Disturbance of delayed match-to-sample in macaques by tetanization of anterior commissure versus limbic system or basal ganglia

Summary Three pig-tailed macaques were trained to select (match) from a pair of colored images that which they had seen (sample) and responded to 5–15 s previously. The anterior commissure (AC) and/or its radiation, various loci in basal ganglia, hippocampal formation and control areas, (splenium of corpus callosum, precentral gyrus, insular cortex), totalling 40 loci, were each tetanized for 4 s during presentation of the sample image, during the delay period, or when the monkey was required to select the matching image. For several loci in the hippocampal formation tetanization at any phase of the task reduced matching to chance levels and gave evidence of electrical after-discharge; but other comparable hippocampal loci had little or no effect. Response to sample or match stimuli were absent during tetanization of basal ganglia or anterior commissure. When finally made, upon cessation of tetanization, responses were equally correct for basal ganglia and control sites, but for AC were at chance levels.     

Effects of sleep deprivation on the activity of selected metabolic enzymes in skeletal muscle

Summary The present study gives the results of a comparison of the recorded and true tibia-calcaneal angles in 17 normal subjects and in 14 patients with abnormally hypoextensible non contracting triceps. 1. For a minimal passive torque, the difference between true and recorded angles varied considerably from one individual to another. The means and ranges for the two groups were respectively: –8 (+7, –21) and –7 (+5, –20). 2. When the passive torque increased as a result of slow passive lengthening of the muscle, the true curve was steeper than the recorded one, owing to differences between the two angle measurements. For each of the two groups the differences in means and ranges were respectively: 6 (0, +13.5) and 8 (3, 12). 3. Subjects made isometric voluntary contractions of the triceps surae at fixed angles which corresponded to step by step muscle lengthening. The resulting true curve was much steeper than the recorded curve. The differences in means and ranges were: 7 (1.5, +15) in children of the two groups and respectively 3 (0, +9) and 12 (10, 14) in adults of the two groups. The present results show that this methodology was the only reliable way of correctly obtaining passive and active torque-angle curves, measuring differences between subjects, appreciating the effects of treatments and these by ascertaining whether or not trophic muscle regulation was defective.