Protoberberine alkaloids from the rhizomes of Corydalis cava were investigated with regard to their influence on the GABA A receptor using radioreceptor assays. Whereas the protoberberine type 2 ...alkaloids, isoapocavidine, corydaline, tetrahydropalmatine, scoulerine and isocorypalmine, increased the specific (3)HBMC-binding in a range of 21 - 49 %, the protoberberine type 1 alkaloids, palmatine, coptisine, dehydroapocavidine, and dehydrocorydaline, had no influence on the binding behaviour of the GABA A receptor. To confirm the modulatory activity of the protoberberine type 2 alkaloids on living cells, GABA A receptor binding studies were performed by fluorescence correlation spectroscopy (FCS) using hippocampal neurons and the fluorescently labelled ligand, muscimol-Alexa (Mu-Alexa). The incubation of hippocampal neurons with 7.5 nM Mu-Alexa showed a specific binding of 5.25 nM (70 %). The evaluation of the autocorrelation curve revealed two different mobilities of receptor ligand complexes, D bound1 = (2.8 +/- 0.91) microm 2/s for the free lateral mobility and D bound2 = (0.14 +/- 0.05) microm 2/s for the hindered mobility. An incubation of hippocampal neurons with 7.5 nM Mu-Alexa and 7.5 nM scoulerine showed a maximal increase of the specific Mu-Alexa binding of approximately 27 % by selectively modulating the amount of receptor-ligand complexes with a hindered mobility (9 % to 27 %).
Abstract
Zolpidem is a hypnotic benzodiazepine site agonist with some γ‐aminobutyric acid (GABA)
A
receptor subtype selectivity. Here, we have tested the effects of zolpidem on the hippocampus of γ2 ...subunit (γ2F77I) point mutant mice. Analysis of forebrain GABA
A
receptor expression with immunocytochemistry, quantitative
3
Hmuscimol and
35
S t‐butylbicyclophosphorothionate (TBPS) autoradiography, membrane binding with
3
Hflunitrazepam and
3
Hmuscimol, and comparison of miniature inhibitory postsynaptic current (mIPSC) parameters did not reveal any differences between homozygous γ2I77/I77 and γ2F77/F77 mice. However, quantitative immunoblot analysis of γ2I77/I77 hippocampi showed some increased levels of γ2, α1, α4 and δ subunits, suggesting that differences between strains may exist in unassembled subunit levels, but not in assembled receptors. Zolpidem (1 µ
m
) enhanced the decay of mIPSCs in CA1 pyramidal cells of control (C57BL/6J, γ2F77/F77) mice by ∼ 60%, and peak amplitude by ∼ 20% at 33–34 °C
in vitro
. The actions of zolpidem (100 n
m
or 1 µ
m
) were substantially reduced in γ2I77/I77 mice, although residual effects included a 9% increase in decay and 5% decrease in peak amplitude. Similar results were observed in CA1 stratum oriens/alveus interneurons. At network level, the effect of zolpidem (10 µ
m
) on carbachol‐induced oscillations in the CA3 area of γ2I77/I77 mice was significantly different compared with controls. Thus, the γ2F77I point mutation virtually abolished the actions of zolpidem on GABA
A
receptors in the hippocampus. However, some residual effects of zolpidem may involve receptors that do not contain the γ2 subunit.
Zolpidem is a hypnotic benzodiazepine site agonist with some gamma -aminobutyric acid (GABA) sub(A) receptor subtype selectivity. Here, we have tested the effects of zolpidem on the hippocampus of ...gamma 2 subunit ( gamma 2F77I) point mutant mice. Analysis of forebrain GABA sub(A) receptor expression with immunocytochemistry, quantitative super(3)Hmuscimol and super(35)S t-butylbicyclophosphorothionate (TBPS) autoradiography, membrane binding with super(3)Hflunitrazepam and super(3)Hmuscimol, and comparison of miniature inhibitory postsynaptic current (mIPSC) parameters did not reveal any differences between homozygous gamma 2I77-I77 and gamma 2F77-F77 mice. However, quantitative immunoblot analysis of gamma 2I77-I77 hippocampi showed some increased levels of gamma 2, alpha 1, alpha 4 and delta subunits, suggesting that differences between strains may exist in unassembled subunit levels, but not in assembled receptors. Zolpidem (1 mu m) enhanced the decay of mIPSCs in CA1 pyramidal cells of control (C57BL-6J, gamma 2F77-F77) mice by similar to 60%, and peak amplitude by similar to 20% at 33-34 degree C in vitro. The actions of zolpidem (100 nm or 1 mu m) were substantially reduced in gamma 2I77-I77 mice, although residual effects included a 9% increase in decay and 5% decrease in peak amplitude. Similar results were observed in CA1 stratum oriens-alveus interneurons. At network level, the effect of zolpidem (10 mu m) on carbachol-induced oscillations in the CA3 area of gamma 2I77-I77 mice was significantly different compared with controls. Thus, the gamma 2F77I point mutation virtually abolished the actions of zolpidem on GABA sub(A) receptors in the hippocampus. However, some residual effects of zolpidem may involve receptors that do not contain the gamma 2 subunit.
Zolpidem is a hypnotic benzodiazepine site agonist with some gamma-aminobutyric acid (GABA)(A) receptor subtype selectivity. Here, we have tested the effects of zolpidem on the hippocampus of gamma2 ...subunit (gamma2F77I) point mutant mice. Analysis of forebrain GABA(A) receptor expression with immunocytochemistry, quantitative (3)Hmuscimol and (35)S t-butylbicyclophosphorothionate (TBPS) autoradiography, membrane binding with (3)Hflunitrazepam and (3)Hmuscimol, and comparison of miniature inhibitory postsynaptic current (mIPSC) parameters did not reveal any differences between homozygous gamma2I77/I77 and gamma2F77/F77 mice. However, quantitative immunoblot analysis of gamma2I77/I77 hippocampi showed some increased levels of gamma2, alpha1, alpha4 and delta subunits, suggesting that differences between strains may exist in unassembled subunit levels, but not in assembled receptors. Zolpidem (1 microm) enhanced the decay of mIPSCs in CA1 pyramidal cells of control (C57BL/6J, gamma2F77/F77) mice by approximately 60%, and peak amplitude by approximately 20% at 33-34 degrees C in vitro. The actions of zolpidem (100 nm or 1 microm) were substantially reduced in gamma2I77/I77 mice, although residual effects included a 9% increase in decay and 5% decrease in peak amplitude. Similar results were observed in CA1 stratum oriens/alveus interneurons. At network level, the effect of zolpidem (10 microm) on carbachol-induced oscillations in the CA3 area of gamma2I77/I77 mice was significantly different compared with controls. Thus, the gamma2F77I point mutation virtually abolished the actions of zolpidem on GABA(A) receptors in the hippocampus. However, some residual effects of zolpidem may involve receptors that do not contain the gamma2 subunit.
Zolpidem is a hypnotic benzodiazepine site agonist with some γ‐aminobutyric acid (GABA)A receptor subtype selectivity. Here, we have tested the effects of zolpidem on the hippocampus of γ2 subunit ...(γ2F77I) point mutant mice. Analysis of forebrain GABAA receptor expression with immunocytochemistry, quantitative 3Hmuscimol and 35S t‐butylbicyclophosphorothionate (TBPS) autoradiography, membrane binding with 3Hflunitrazepam and 3Hmuscimol, and comparison of miniature inhibitory postsynaptic current (mIPSC) parameters did not reveal any differences between homozygous γ2I77/I77 and γ2F77/F77 mice. However, quantitative immunoblot analysis of γ2I77/I77 hippocampi showed some increased levels of γ2, α1, α4 and δ subunits, suggesting that differences between strains may exist in unassembled subunit levels, but not in assembled receptors. Zolpidem (1 µm) enhanced the decay of mIPSCs in CA1 pyramidal cells of control (C57BL/6J, γ2F77/F77) mice by ∼ 60%, and peak amplitude by ∼ 20% at 33–34 °C in vitro. The actions of zolpidem (100 nm or 1 µm) were substantially reduced in γ2I77/I77 mice, although residual effects included a 9% increase in decay and 5% decrease in peak amplitude. Similar results were observed in CA1 stratum oriens/alveus interneurons. At network level, the effect of zolpidem (10 µm) on carbachol‐induced oscillations in the CA3 area of γ2I77/I77 mice was significantly different compared with controls. Thus, the γ2F77I point mutation virtually abolished the actions of zolpidem on GABAA receptors in the hippocampus. However, some residual effects of zolpidem may involve receptors that do not contain the γ2 subunit.
Agonists of the allosteric benzodiazepine site of GABA
A receptors bind at the interface of the α and γ subunits. Here, we tested the in vivo contribution of the γ2 subunit to the actions of ...zolpidem, an α1 subunit selective benzodiazepine agonist, by generating mice with a phenylalanine (F) to isoleucine (I) substitution at position 77 in the γ2 subunit. The γ2F77I mutation has no major effect on the expression of GABA
A receptor subunits in the cerebellum. The potency of zolpidem, but not that of flurazepam, for the inhibition of
3Hflunitrazepam binding to cerebellar membranes is greatly reduced in γ2I77/I77 mice. Zolpidem (1 μM) increased both the amplitude and decay of miniature inhibitory postsynaptic currents (mIPSCs) in Purkinje cells of control C57BL/6 (34% and 92%, respectively) and γ2F77/F77 (20% and 84%) mice, but not in those of γ2F77I mice. Zolpidem tartrate had no effect on exploratory activity (staircase test) or motor performance (rotarod test) in γ2I77/I77 mice at doses up to 30 mg/kg (i.p.) that strongly sedated or impaired the control mice. Flurazepam was equally effective in enhancing mIPSCs and disrupting performance in the rotarod test in control and γ2I77/I77 mice. These results show that the effect of zolpidem, but not flurazepam, is selectively eliminated in the brain by the γ2F77I point mutation.
Agonists of the allosteric benzodiazepine site of GABA sub(A) receptors bind at the interface of the alpha and gamma subunits. Here, we tested the in vivo contribution of the gamma 2 subunit to the ...actions of zolpidem, an alpha 1 subunit selective benzodiazepine agonist, by generating mice with a phenylalanine (F) to isoleucine (I) substitution at position 77 in the gamma 2 subunit. The gamma 2F77I mutation has no major effect on the expression of GABA sub(A) receptor subunits in the cerebellum. The potency of zolpidem, but not that of flurazepam, for the inhibition of super(3)Hflunitrazepam binding to cerebellar membranes is greatly reduced in gamma 2I77/I77 mice. Zolpidem (1 mu M) increased both the amplitude and decay of miniature inhibitory postsynaptic currents (mIPSCs) in Purkinje cells of control C57BL/6 (34% and 92%, respectively) and gamma 2F77/F77 (20% and 84%) mice, but not in those of gamma 2F77I mice. Zolpidem tartrate had no effect on exploratory activity (staircase test) or motor performance (rotarod test) in gamma 2I77/I77 mice at doses up to 30 mg/kg (i.p.) that strongly sedated or impaired the control mice. Flurazepam was equally effective in enhancing mIPSCs and disrupting performance in the rotarod test in control and gamma 2I77/I77 mice. These results show that the effect of zolpidem, but not flurazepam, is selectively eliminated in the brain by the gamma 2F77I point mutation.
Agonists of the allosteric benzodiazepine site of GABAA receptors bind at the interface of the alpha and gamma subunits. Here, we tested the in vivo contribution of the gamma2 subunit to the actions ...of zolpidem, an alpha1 subunit selective benzodiazepine agonist, by generating mice with a phenylalanine (F) to isoleucine (I) substitution at position 77 in the gamma2 subunit. The gamma2F77I mutation has no major effect on the expression of GABAA receptor subunits in the cerebellum. The potency of zolpidem, but not that of flurazepam, for the inhibition of 3Hflunitrazepam binding to cerebellar membranes is greatly reduced in gamma2I77/I77 mice. Zolpidem (1 microM) increased both the amplitude and decay of miniature inhibitory postsynaptic currents (mIPSCs) in Purkinje cells of control C57BL/6 (34% and 92%, respectively) and gamma2F77/F77 (20% and 84%) mice, but not in those of gamma2F77I mice. Zolpidem tartrate had no effect on exploratory activity (staircase test) or motor performance (rotarod test) in gamma2I77/I77 mice at doses up to 30 mg/kg (i.p.) that strongly sedated or impaired the control mice. Flurazepam was equally effective in enhancing mIPSCs and disrupting performance in the rotarod test in control and gamma2I77/I77 mice. These results show that the effect of zolpidem, but not flurazepam, is selectively eliminated in the brain by the gamma2F77I point mutation.
Health-related quality of life (HRQoL) comprises different domains of physical, mental, and social well-being. In this analysis, we focus on sexual quality of life in Hodgkin Lymphoma (HL) patients.
...Four-thousand one-hundred and sixty patients enroled in the HD10-HD12 trials underwent HRQoL assessment. Instruments included the Quality of Life Questionnaire for survivors (QLQ-S), combining the European Organisation for Research and Treatment of Cancer QLQ-C30, Multidimensional fatigue (FA) inventory (MFI-20) and an additional sexual functioning (SX) scale. We describe SX up to 27 months after therapy and analyse relationship to stage, age, gender, FA, social functioning, and therapy. Statistical methods range from descriptive statistics to a classification of SX courses, and a longitudinal structural equations model with full information maximum likelihood estimation of missing data. In the analysis, a score below 50 was used to describe severe sexual dysfunction.
Three-thousand two-hundred and eight patients provided data on SX. Patients in advanced stages reported lower SX than patients in early stages both, before and after the treatment. During follow-up, an improvement of SX compared with baseline was detected, except for those ≥50 years. Patients in early stages reached normal SX, whereas advanced-stage patients remained below the reference value for healthy controls. Sexual functioning during follow-up was significantly and strongly related to previous SX, other HRQoL measures, age, and stage, and to lesser degree with gender and chemotherapy.
Overall, HL patients have a decreased sexual quality of life at baseline, which improves after therapy and normalises in early-stage patients. Importantly, long-term SX is more closely related to patient characteristics and SX at baseline than to the intensity of treatment.
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