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  • 1
    Electronic Resource
    Electronic Resource
    Lichtmikroskopische Dünnschnitt-Technik an der Haut (1966)
    Springer
    Archives of dermatological research 224 (1966), S. 416-423 
    Details
    ISSN: 1432-069X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Description / Table of Contents: Zusammenfassung Es wird über die Anwendungsmöglichkeit der elektronen mikroskopischen Einbettungs- und Dünnschnitt-Technik für lichtmikroskopische Untersuchungen an Hautschnitten (2,0–3,0 μm) berichtet.
    Notes: Summary Thin sections (2–3 μm) can easily be obtained from skin fixed in osmium and embedded in Polyester (Vestopal). Such sections can also be stained by several histological and histochemical routine procedures for light microscopical observations (Table 2).
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00518307
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  • 2
    Electronic Resource
    Electronic Resource
    Entwicklungsfragen bei Apfelgehölzen (1954)
    Springer
    Theoretical and applied genetics 24 (1954), S. 346-352 
    Details
    ISSN: 1432-2242
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Zusammenfassung Die mitgeteilten Ergebnisse lassen erkennen, daß das Entwicklungsgeschehen bei Apfelsämlingen weitgehend von der J.-Th. abweicht. Das plötzliche Umschlagen des ganzen Sämlings sowie die Kopplung der Fertilität an diesen Umschlag entspricht nicht den Tatsachen. Eine Zweiteilung der Sterilität in eine absolute und eine ernährungsphysiologisch bedingte verliert ihren Wert, nachdem bewiesen ist, daß Dorntriebe, also angeblich absolut sterile Organe, blühen können. Die Beschleunigung des Stadienablaufes durch äußere Einflüsse ist möglich. Eine unbedingte Konstanz der sogenannten “Jugend-” und “Altersformzone” ist nicht gegeben. Das Verhalten des Typ IX widerspricht den Forderungen der J.-Th. insofern, als er sich weder wie eine “Jugendform” noch wie eine “Altersform” verhält, sondern die Kennzeichen beider Entwicklungsstadien gleichzeitig trägt. Die Kenntnis des Entwicklungsgeschehens steht noch viel zu sehr im Aufang, um ein derart abgerundetes Bild anbieten zu können, wie es die J.-Th. tut. Bei der langsamen Entwicklung der Obstgehölze ist eine so kurzfristige Beobachtungszeit, wie sie sich die Vertreter der J.-Th. für die Aufstellung ihrer Lehre gestattet haben, ungenügend.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00710394
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  • 3
    Electronic Resource
    Electronic Resource
    Regulation of the Na+2Cl–K+ cotransporter in in vitro perfused rectal gland tubules of Squalus acanthias (1998)
    Springer
    Pflügers Archiv 436 (1998), S. 521-528 
    Details
    ISSN: 1432-2013
    Keywords: Key words cAMP ; Cl ; channels ; Cl ; secretion ; Exocrine secretion ; K+ channels ; Volume regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  Previously it has been shown that the Na+2Cl–K+ cotransporter accepts NH4 + at its K+ binding site. This property can be used to estimate its transport rates by adding NH4 + to the bath and measuring the initial furosemide-dependent rates of change in BCECF fluorescence. We have utilized this technique to determine the regulation of the furosemide-inhibitable Na+2Cl–K+ cotransporter in in vitroperfused rectal gland tubules (RGT) of Squalus acanthias. Addition of NH4 + to the bath (20 mmol/l) led to an initial alkalinization, corresponding to NH3 uptake. This was followed by an acidification, corresponding to NH4 + uptake. The rate of this uptake was quantified by exponential curve fitting and is given in arbitrary units (Δfluorescence/time). This acidification could be completely inhibited by furosemide. In the absence of any secretagogue preincubation of RGT in a low Cl– solution (6 mmol/l, low Cl–) for 10 min enhanced the uptake rate significantly from 4.04±0.51 to 12.7±1.30 (n=5). The addition of urea (200 mmol/l) was without effect, but the addition of 300 mmol/l mannitol (+300 mannitol) enhanced the rate significantly from 7.24±1.33 to 14.7±4.6 (n=6). Stimulation of NaCl secretion by a solution maximizing the cytosolic cAMP concentration (Stim) led to a significant increase in NH4 + uptake rate from 5.00±1.33 to 13.3±1.54 (n=6). Similar results were obtained in the additional presence of Ba2+ (1 mmol/l): the uptake rate was increased significantly from 4.23±0.34 to 15.1±1.86 (n=16). In the presence of Stim low Cl– had no additional effect on the uptake rate: 15.1±3.1 versus 15.2±2.8 in high Cl– (n=6). The uptake rate in Stim containing additional +300 mannitol (22.3±4.0, n=5) was not significantly different from that obtained with Stim or +300 mannitol alone. By whatever mechanism the NH4 + uptake rate was increased furosemide (500 µmol/l) always reduced this rate to control values. Hence three manoeuvres enhanced furosemide-inhibitable uptake rates of the Na+2Cl–K+ cotransporter probably independently: (1) lowering of cytosolic Cl– concentration; (2) cell shrinkage; and (3) activation by cAMP.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050667
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  • 4
    Electronic Resource
    Electronic Resource
    pH-regulatory mechanisms in in vitro perfused rectal gland tubules of Squalus acanthias (1998)
    Springer
    Pflügers Archiv 436 (1998), S. 248-254 
    Details
    ISSN: 1432-2013
    Keywords: Key words Cl ; secretion ; DIDS ; HCO3 ; /Cl ; exchange ; HOE694 ; Na+/H+ exchange
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  Isolated in vitro perfused rectal gland tubules (RGT) were preincubated with the pH-sensitive dye 2′,7′-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) and pH-regulatory mechanisms were studied. A reduction of bath Cl– concentration from 269 to 6 mmol/l increased the fluorescence ratio 488/436 [corresponding to cytosolic pH (pHi)] slightly but significantly (n=10). Depolarization by Ba2+ (1 mmol/l) or a bath solution containing 30 mmol/l K+ (n=4–6) increased the fluorescence ratio (pHi). These data suggest that HCO3 – uptake and/or H+ extrusion is dependent on Cl– and/or voltage. A reduction of bath Na+ from 278 to 5 mmol/l reduced the ratio significantly (n=3). Addition of trimethylamine (Trima+, 20 mmol/l) alkalinized cytosolic pH (n=7). Similarly, addition of NH4 + (20 mmol/l) led to an initial alkalinization and a strong acidification when NH4 + was removed (n=59). The initial pHi-recovery rates after NH4 + removal were quantified and the responsible H+ extrusion and/or HCO3 – import systems were examined. The recovery was almost completely abolished when the extracellular Na+ concentration was reduced to 5 mmol/l. In the presence of normal Na+, recovery was slower in the absence as compared to the presence of HCO3 – (n=5). It was inhibited by 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS) (0.5 mmol/l, n=11) in the presence of HCO3 – and in the absence of HCO3 – by the Na+/H+-exchange blocker HOE694 (0.5 mmol/l, n=6). These data suggest that acid extrusion probably occurs by basolateral Na+-2HCO3 –/Cl– exchange in the presence of HCO3 – and by basolateral Na+/H+ exchange in the absence of HCO3 –. Luminal perfusion with a solution containing a low Cl– concentration (6 mmol/l) increased the fluorescence ratio (pHi) (n=5). The ratio (pHi) was further increased and pH recovery further delayed by basolateral addition of Trima+ (20 mmol/l, n=3). These data suggest that the HCO3 –/Cl– exchanger is present in the luminal membrane. Luminal HCO3 –/Cl– exchange and basolateral Na+-2HCO3 –/Cl– exchange may work in tandem to secrete HCO3 – and exchange it for luminal Cl–.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050629
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  • 5
    Electronic Resource
    Electronic Resource
    Does stimulation of NaCl secretion in in vitro perfused rectal gland tubules of Squalus acanthias increase membrane capacitance? (1998)
    Springer
    Pflügers Archiv 436 (1998), S. 538-544 
    Details
    ISSN: 1432-2013
    Keywords: Key words CFTR ; Cl ; channels ; Cl ; secretion ; Endocytosis ; Exocrine secretion ; Exocytosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  NaCl secretion in rectal gland tubules (RGT) of Squalus acanthias requires the activation of Cl– channels in the luminal membrane. The RGT and its mechanism of activation are an early evolutionary paradigm of exocrine secretion. The respective Cl– channels probably resemble the shark equivalent of the cystic fibrosis transmembrane conductance regulator (CFTR). Activation of these Cl– channels occurs via cAMP. It has been hypothesized that the activation of CFTR occurs via exocytosis or inhibited endocytosis. To examine this question directly by electrical measurements we have performed whole-cell patch-clamp analyses of in vitro perfused RGT. NaCl secretion was stimulated by a solution (Stim) containing forskolin (10 µmol/l), dibutyryl-cAMP (0.5 mmol/l) and adenosine (0.5 mmol/l). This led to the expected strong depolarization and an increase in membrane conductance (G m). The membrane capacitance (C m) was measured by a newly devised two-frequency synchronous detector method. It was increased by Stim significantly from 5.00±0.22 to 5.17±0.21 pF (n=50). The increase in C m correlated with the increase in G m with a slope of 51 fF/nS. Next the effect of furosemide (500 µmol/l) was examined in previously stimulated RGT. Furosemide was supposed to inhibit coupled Na+2Cl–K+ uptake and to reduce cell volume but not membrane trafficking of Cl– channels. Furosemide reduced G m slightly (due to the fall in cytosolic Cl– concentration) and C m to the same extent by which Stim had increased it. Both changes were statistically significant, and the slope of ΔC m/ΔG m was similar to that caused by Stim. Inhibitors of microtubules or actin (colchicine, phalloidin and cytochalasin D added at 10 µmol/l to the pipette solution and dialysed for 〉10 min) did not alter cell voltage, G m or C m, nor did these inhibitors abolish the stimulatory effect of cAMP. These data suggest that the small C m changes observed with Stim reflect a minor cell volume increase and an ”unfolding” of the plasma membrane. The present data do not support the exocytosis/endocytosis hypothesis of cAMP-mediated activation of Cl– channels in these cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050669
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  • 6
    Electronic Resource
    Electronic Resource
    The role of exocytosis in the activation of the chloride conductance in Chinese hamster ovary cells (CHO) stably expressing CFTR (1997)
    Springer
    Pflügers Archiv 434 (1997), S. 779-784 
    Details
    ISSN: 1432-2013
    Keywords: Keywords Cystic fibrosis ; Exocytosis ; Cl ; channel ; Ca2+ activated Cl ; channel ; CFTR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  The aim of this study was to examine the question of whether activation of wt-CFTR (wild-type cystic fibrosis transmembrane conductance regulator) by cAMP and the opening of a Cl–conductance is paralleled by exocytosis and corresponding increases in membrane capacitance. To this end three types of Chinese hamster ovary (CHO) cells were examined: a control group of CHO cells; a group of CHO cells stably expressing wt-CFTR at high levels (also called BQ2-CHO); and a group of CHO cells stably expressing the frequent mutation ΔF508-CFTR. Whole-cell patch-clamp studies were performed to measure the membrane voltage (V m), the membrane conductance (G m) and the membrane capacitance (C m). C m was assessed by a two-frequency lock-in amplifier method. Forskolin (Fsk, 0.1 μmol/l) and isobutylmethylxanthine (IBMX, 0.1 mmol/l) were used to increase cytosolic cAMP. It is shown that Fsk and IBMX had no effect on V m and G m in control CHO and ΔF508-CFTR-CHO cells. Fsk and IBMX depolarized wt-CFTR-expressing CHO cells significantly (from –40 ± 1.5 to –32 ± 1.6 mV, n = 41) and enhanced G m strongly from 5.0 ± 0.9 to 36 ± 3.9 nS (n = 65). The conductance increase was mostly for Cl–, because under stimulated conditions a reduction in bath Cl–concentration depolarized these cells further and significantly from –26 ± 1.8 to –10 ± 1.2 mV (n = 16). This conductance had the characteristic wt-CFTR selectivity of Br– 〉 Cl– 〉 I–(n = 16). Despite this large increase in the Fsk- and IBMX-induced conductance C m was not altered significantly (15.5 versus 15.7 pF, n = 50). These data indicate that stable overexpression of wt-CFTR but not of ΔF508-CFTR in CHO cells induces a cAMP-activated Cl–conductance. The activation of this large conductance obviously proceeds with little if any exocytosis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050465
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  • 7
    Electronic Resource
    Electronic Resource
    The role of cytosolic Ca2+ in the secretion of NaCl in isolated in vitro perfused rectal gland tubules of Squalus acanthias (1998)
    Springer
    Pflügers Archiv 436 (1998), S. 133-140 
    Details
    ISSN: 1432-2013
    Keywords: Key words Cl ; secretion ; Cl ; channels ; K+ channels ; cGMP ; cAMP ; Cytosolic Ca2+ ; Exocrine secretion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  In many exocrine glands cytosolic Ca2+ ([Ca2+]i) plays a pivotal role in stimulation-secretion coupling. In the rectal gland of the dogfish Squalus acanthias this appears not to be the case and it is believed that secretion is mainly controlled by the Cl– conductance of the luminal membrane. We have examined this question in a study of isolated in vitro perfused rectal gland tubules (RGT). Three types of measurements were performed: (1) measurements of [Ca2+]i by the fura-2 technique; (2) measurements of transepithelial electrical parameters, i.e. transepithelial voltage (V te), transepithelial resistance (R te), the equivalent short-circuit current (I sc) and the voltage across the basolateral membrane (V bl), and (3) whole-cell patch-clamp measurements of cellular voltage (V m), conductance (G m) and membrane capacitance (C m). The data indicates that carbachol (CCH) increases [Ca2+]i by increasing store release and Ca2+ influx. Other agonists, producing cytosolic cAMP, also increased [Ca2+] by enhancing Ca2+ influx. CCH hyperpolarized these cells and enhanced G m significantly. The effect of CCH on V te and I sc was most marked under control conditions and disappeared in RGT otherwise stimulated by agonists that lead to cAMP production. It is concluded that [Ca2+]i plays a major role in the stimulation of NaCl secretion in RGT by enhancing the basolateral K+ conductance. cAMP-producing agonists enhance [Ca2+]i by increased Ca2+ influx. CCH releases Ca2+ from respective stores. CCH, unlike the cAMP-producing agonists, only increases basolateral K+ conductance. It modulates secretion especially under conditions in which the cAMP pathway is not fully activated.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050614
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  • 8
    Electronic Resource
    Electronic Resource
    The cellular mechanisms of Cl– secretion induced by C-type natriuretic peptide (CNP). Experiments on isolated in vitro perfused rectal gland tubules of Squalus acanthias (1999)
    Springer
    Pflügers Archiv 438 (1999), S. 15-22 
    Details
    ISSN: 1432-2013
    Keywords: Key words cGMP ; Cl ; secretion ; C-type natriuretic peptide ; NaCl secretion ; Squalus acanthia
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  We have examined the mechanism whereby C-type natriuretic peptide (CNP), an agonist acting through the second messenger cGMP, enhances NaCl secretion in the rectal gland of Squalus acanthias. Single rectal gland tubules (RGT) were dissected manually, perfused in vitro and equivalent short-circuit current [I sc=transepithelial voltage/transepithelial resistance (R te)] as well as basolateral membrane voltage (V bl) were measured. CNP was added to luminal and basolateral perfusates at concentrations between 1 and 1000 nmol/l and its effects on the above parameters were compared to those of a ”stimulation cocktail” (Stim, containing dibutyryl cAMP, adenosine and forskolin) that maximally enhances cytosolic cAMP, and other agonists and hormones such as guanylin, vasoactive intestinal peptide (VIP), and adenosine. CNP had no effect from the luminal side (n=6). Its effects from the basolateral side consisted of a substantial increase in I sc (–31.6±7.7 to –316±82.2 µA/cm2, n=15). CNP significantly depolarized the luminal membrane from –87.4±1.0 to –82.3±2.6 mV (n=12). V bl was not changed (n=12) but the fractional conductance for K+ was increased (n=3). These effects were qualitatively and even quantitatively comparable to those of other agonists acting via cytosolic cAMP, but were less marked than those caused by Stim (n=64). The effects of VIP and CNP on I sc were not additive (n=5). The cytosolic Ca2+ concentration ([Ca2+]i) was monitored using the fura-2 fluorescence ratio (FFR 340/380 nm) and it was found that CNP, like agonists acting via cAMP, enhances FFR significantly from 1.02±0.05 to 1.32±0.05 (n=8) with a time constant in the 1–2 min in range. Our data suggest that CNP, acting via the second messenger cGMP, induces a marked increase in I sc in the rectal gland. The concomitant fall in R te corresponds to increases in the luminal membrane Cl– conductance and in the basolateral membrane K+ conductance. The latter effect is probably due to an increase in [Ca2+]i.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050874
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