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Biomedical subjects

Volker Hagen

Publications and source records attributed to Volker Hagen.

11 recordsLinked to original sources
PubMed2003

A sperm-activating peptide controls a cGMP-signaling pathway in starfish sperm.

Peptides released from eggs of marine invertebrates play a central role in fertilization. About 80 different peptides from various phyla have been isolated, however, with one exception, their respective receptors on the sperm surface have not been unequivocally identified and the pertinent signaling pathways remain ill defined. Using rapid mixing techniques and novel membrane-permeable caged compounds of cyclic nucleotides, we show that the sperm-activating peptide asterosap evokes a fast and transient increase of the cGMP concentration in sperm of the starfish Asterias amurensis, followed by a transient cGMP-stimulated increase in the Ca(2+) concentration. In contrast, cAMP levels did not change significantly and the Ca(2+) response evoked by photolysis of caged cAMP was significantly smaller than that using caged cGMP. By cloning of cDNA and chemical crosslinking, we identified a receptor-type guanylyl cyclase in the sperm flagellum as the asterosap-binding protein. Sperm respond exquisitely sensitive to picomolar concentrations of asterosap, suggesting that the peptide serves a chemosensory function like resact, a peptide involved in chemotaxis of sperm of the sea urchin Arbacia punctulata. A unifying principle emerges that chemosensory transduction in sperm of marine invertebrates uses cGMP as the primary messenger, although there may be variations in the detail.

Amino Acid Sequence↗
PubMed2003

[7-(Dialkylamino)coumarin-4-yl]methyl-Caged Compounds as Ultrafast and Effective Long-Wavelength Phototriggers of 8-Bromo-Substituted Cyclic Nucleotides.

[7-(Dimethylamino)coumarin-4-yl]methyl (DMACM) and [7-(diethylamino)coumarin-4-yl]methyl (DEACM) esters of 8-bromoadenosine 3',5'-cyclic monophosphate (8-Br-cAMP) and 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP) are described as novel caged compounds for 8-bromo-substituted cyclic nucleotides. Synthesis is accomplished by treatment of the free acids of the cyclic nucleotides with the corresponding 7(dialkylamino)-substituted 4(diazomethyl)coumarins. Irradiation of the DMACM- and DEACM-caged cyclic nucleotides with UV light stimulates the release of the cyclic nucleotides within roughly a nanosecond. The new caged compounds are resistant to hydrolysis in aqueous buffers and exhibit long-wavelength absorption properties with maxima at 400 nm, high extinction coefficients, and high quantum yields (0.15-0.31). Their favorable properties render these compounds the most efficient and rapid phototriggers of 8-bromo-substituted cyclic nucleotides known. The usefulness of the compounds for physiological studies under nondamaging light conditions was examined in HEK293 cells expressing the alpha subunit of the cyclic-nucleotide-gated (CNG) channel of cone photoreceptors (CNGA3) and of olfactory neurons (CNGA2) by using confocal laser scanning microscopy and the patch clamp technique.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2003

DMACM-caged adenosine nucleotides: ultrafast phototriggers for ATP, ADP, and AMP activated by long-wavelength irradiation.

The development of new photocleavable adenosine nucleotides based on the photochemistry of [7-(dimethylamino)coumarin-4-yl]methyl (DMACM) esters is described. The phototriggers liberate adenosine triphosphate (ATP), diphosphate, and monophosphate upon UV/Vis irradiation between 334 and 405 nm. The efficiency of photocleavage at long wavelengths is high as a result of a combination of appropriate quantum yields and intensive absorptivities. By using time-resolved fluorescence spectroscopy, we determined a lower limit of 1.6 x 10(9) s(-1) for the rate constant of the release of ATP from DMACM-caged ATP. The favorable properties of DMACM-caged ATP were confirmed in physiological studies by confocal laser scanning microscopy. We were able to uncage DMACM-caged ATP in cultures of mouse astrocytes and in brain tissue slices from mice and were also able to measure the effect of photoreleased ATP on the cellular response of astrocytes, namely the ability of the ATP to evoke Ca(2+) ion waves.

Adenosine Diphosphate↗
PubMed2003

Structural proton diffusion along lipid bilayers.

For H(+) transport between protein pumps, lateral diffusion along membrane surfaces represents the most efficient pathway. Along lipid bilayers, we measured a diffusion coefficient of 5.8 x 10(-5) cm(2) s(-1). It is too large to be accounted for by vehicle diffusion, considering proton transport by acid carriers. Such a speed of migration is accomplished only by the Grotthuss mechanism involving the chemical exchange of hydrogen nuclei between hydrogen-bonded water molecules on the membrane surface, and the subsequent reorganization of the hydrogen-bonded network. Reconstitution of H(+)-binding sites on the membrane surface decreased the velocity of H(+) diffusion. In the absence of immobile buffers, structural (Grotthuss) diffusion occurred over a distance of 100 micro m as shown by microelectrode aided measurements of the spatial proton distribution in the immediate membrane vicinity and spatially resolved fluorescence measurements of interfacial pH. The efficiency of the anomalously fast lateral diffusion decreased gradually with an increase in mobile buffer concentration suggesting that structural diffusion is physiologically important for distances of approximately 10 nm.

Diffusion↗
PubMed2003

The signal flow and motor response controling chemotaxis of sea urchin sperm.

The signalling pathway and the behavioural strategy underlying chemotaxis of sperm are poorly understood. We have studied the cellular events and motor responses that mediate chemotaxis of sperm from the sea urchin Arbacia punctulata. Here we show that resact, a chemoattractant peptide, initiates a rapid and transient rise in the concentration of cyclic GMP, followed by a transient influx of Ca2+. The binding of a single resact molecule elicits a Ca2+ response, and 50-100 bound molecules saturate the response. The ability to register single molecules is reminiscent of the single-photon sensitivity of rod photoreceptors. Both resact and cyclic nucleotides cause a turn or brief tumbling in the swimming path of sperm. We conclude that a cGMP-mediated increase in the Ca2+ concentration induces the primary motor response of sperm to the chemoattractant.

Animals↗
PubMed2003

Cyclic AMP is sufficient for triggering the exocytic recruitment of aquaporin-2 in renal epithelial cells.

The initial response of renal epithelial cells to the antidiuretic hormone arginine vasopressin (AVP) is an increase in cyclic AMP. By applying immunofluorescence, cell membrane capacitance and transepithelial water flux measurements we show that cAMP alone is sufficient to elicit the antidiuretic cellular response in primary cultured epithelial cells from renal inner medulla, namely the transport of aquaporin-2 (AQP2)-bearing vesicles to, and their subsequent fusion with, the plasma membrane (AQP2 shuttle). The AQP2 shuttle is evoked neither by AVP-independent Ca(2+) increases nor by AVP-induced Ca(2+) increases. However, clamping cytosolic Ca(2+) concentrations below resting levels at 25 nM inhibited exocytosis. Exocytosis was confined to a slow monophasic response, and readily releasable vesicles were missing. Analysis of endocytic capacitance steps revealed that cAMP does not decelerate the retrieval of AQP2 from the plasma membrane. Our data suggest that cAMP initiates an early step, namely the transport of AQP2-bearing vesicles towards the plasma membrane, and do not support a regulatory function for Ca(2+) in the AQP2 shuttle.

Animals↗
PubMed2002

Deactivation behavior and excited-state properties of (coumarin-4-yl)methyl derivatives. 2. Photocleavage of selected (coumarin-4-yl)methyl-caged adenosine cyclic 3',5'-monophosphates with fluorescence enhancement.

A series of axial and equatorial diastereomers of (coumarin-4-yl)methyl-caged adenosine cyclic 3',5'-monophosphates (cAMPs), 1-6, having methoxy, dialkylamino, or no substituent in the 6- and/or 7-positions, and their corresponding 4-(hydroxymethyl)coumarin photoproducts 7-12 have been synthesized. The photochemical and UV/vis spectroscopical properties (absorption and fluorescence) of 1-6 and 7-12 have been examined in methanol/aqueous HEPES buffer solution. Donor substitution in the 6-position causes a strong bathochromic shift of the long-wavelength absorption band, whereas substitution in the 7-position leads only to a weak red shift. The photochemical cleavage of the caged cAMPs was investigated, and the photoproducts were analyzed. Photochemical quantum yields, fluorescence quantum yields, and lifetimes of the excited singlet states were determined. The highest values of photochemical quantum yields (photo-S(N)1 mechanism) were obtained with caged cAMPs having a donor substituent in the 7-position of the coumarin moiety, caused by electronic stabilization of the intermediately formed coumarinylmethyl cation. With donor substitution in the 6-position, the resulting moderate electronic stabilization of the coumarinylmethyl cation is overcompensated by the strong bathochromic shift, reducing the energy gap between the excited-state S(1) and the corresponding coumarinylmethyl cation. The rate constant for the ester cleavage and liberation of cAMP is about 10(9) s(-1), estimated for the axial isomer of 6 by analysis of the fluorescence increase of the alcohol 12 formed upon laser pulse photolysis.

Coumarins↗
PubMed2002

Rapid photolytic release of cytidine 5'-diphosphate from a coumarin derivative: a new tool for the investigation of ribonucleotide reductases.

In order to study the long-range radical transfer in the Escherichia coli ribonucleotide reductase (RNR), caged cytidine 5'-diphosphate (CDP) 1 was synthesized, which contains the photolabile (7-diethylaminocoumarin-4-yl)methyl moiety. The caged CDP 1 triggers the release of CDP when irradiated at wavelengths between 365 and 436 nm. The rate constant of the formation of alcohol 2 and cytidine 5'-diphosphate 3 is 2x10(8) s(-1) and the quantum efficiency for the disappearance of caged CDP 1 is 2.9%.

Coumarins↗
PubMed2002

Imatinib mesylate therapy in patients with gastrointestinal stromal tumors and impaired liver function.

Hepatic and peritoneal metastases are the most frequent metastatic lesions in patients with gastrointestinal stromal tumors (GIST), and may result in intra- or extrahepatic cholestasis and altered drug metabolism. While the tyrosine kinase inhibitor imatinib, which has been recently shown to represent the treatment of choice for GIST, is primarily metabolized by the liver, data on the pharmacokinetics and the tolerability of imatinib in patients with increased cholestasis parameters are not yet available. We here report on two patients who received imatinib in the presence of increased bilirubin and/or cholestasis parameters. With a follow-up duration of 3-4 months, we observed no toxicities outside of well-known side effects including some degree of myelosuppression and fluid retention. This report may aid in the decision of imatinib being given under close surveillance to this kind of patients.

Aged↗