A program of stringently-regulated gene expression is thought to be a fundamental component of th... more A program of stringently-regulated gene expression is thought to be a fundamental component of the circadian clock. Although recent work has implicated a role for E-box-dependent transcription in circadian rhythmicity, the contribution of other enhancer elements has yet to be assessed. Here, we report that cells of the suprachiasmatic nuclei (SCN) exhibit a prominent circadian oscillation in cAMP response element (CRE)-mediated gene expression. Maximal reporter gene expression occurred from late-subjective night to mid-subjective day. Cycling of CRE-dependent transcription was not observed in other brain regions, including the supraoptic nucleus and piriform cortex. Levels of the phospho-active form of the transcription factor CREB (P-CREB) varied as a function of circadian time. Peak P-CREB levels occurred during the mid-to late-subjective night. Furthermore, photic stimulation during the subjective night, but not during the subjective day, triggered a marked increase in CRE-mediated gene expression in the SCN. Reporter gene experiments showed that activation of the p44/42 mitogen-activated protein kinase signaling cascade is required for Ca 2؉-dependent stimulation of CRE-mediated transcription in the SCN. These findings reveal the CREB/CRE transcriptional pathway to be circadian-regulated within the SCN, and raise the possibility that this pathway provides signaling information essential for normal clock function.
The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in ... more The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in intact coho salmon follicles and isolated follicular layers at various stages of oocyte maturation, from late vitellogenesis until the completion of germinal vesicle breakdown (GVBD). In granulosa layers from all stages, LH, but not FSH, stimulated 17␣,20-dihydroxy-4-pregnen-3-one (17,20-P) production. In theca-interstitial layers from all stages, FSH and LH stimulated steroid production, LH being more potent than FSH. The basal steroid output of intact follicles was significantly lower than that of isolated follicular layers, and their response to FSH and LH also differed. First, the intact follicles produced 17␣-hydroxyprogesterone in response to FSH during the central germinal vesicle stage while thecainterstitial layers did not. Second, estradiol-17 production was not inhibited by LH during final oocyte maturation in intact follicles, as observed for granulosa layers. Our results indicate that LH is the determining factor regulating the production of the maturation-inducing steroid, 17,20-P, and the induction of GVBD in the salmonid ovary. In summary, we have provided evidence for maturation-associated changes in the effects of FSH and LH in the salmonid ovary, which further supports the hypothesis that FSH and LH have distinct functions in the teleost ovary.
Specific subcellular targeting and spatial arrangement of signaling molecules are important for e... more Specific subcellular targeting and spatial arrangement of signaling molecules are important for efficient signal transduction. The neuro-specific type-I adenylyl cyclase (AC1) is stimulated by Ca 2+ , and plays an essential role in neurodevelopment and neuroplasticity. We generated hemagglutinin (HA)-tagged AC1 to study its subcellular localization in cultured neurons. The HA-tagged AC1 has similar enzymatic activity and regulatory properties to that of non-tagged protein. HA-AC1 targeted to both apical and basolateral domains in the epithelial Madin-Darby canine kidney (MDCK) cells, and it was found in both axons and dendrites in cultured hippocampal neurons as well as in cerebellar granule neurons. Interestingly, AC1 showed a distinct punctate form of immunostaining in MDCK cells and transfected neurons, suggesting it targets to specific subcellular domains. By immunostaining with different synaptic markers, we found that AC1 puncta were located at the excitatory synapses in cerebellar granule neurons. Our data provide a possible cellular mechanism for the physiological role of AC1 in neuroplasticity.
A program of stringently-regulated gene expression is thought to be a fundamental component of th... more A program of stringently-regulated gene expression is thought to be a fundamental component of the circadian clock. Although recent work has implicated a role for E-box-dependent transcription in circadian rhythmicity, the contribution of other enhancer elements has yet to be assessed. Here, we report that cells of the suprachiasmatic nuclei (SCN) exhibit a prominent circadian oscillation in cAMP response element (CRE)-mediated gene expression. Maximal reporter gene expression occurred from late-subjective night to mid-subjective day. Cycling of CRE-dependent transcription was not observed in other brain regions, including the supraoptic nucleus and piriform cortex. Levels of the phospho-active form of the transcription factor CREB (P-CREB) varied as a function of circadian time. Peak P-CREB levels occurred during the mid-to late-subjective night. Furthermore, photic stimulation during the subjective night, but not during the subjective day, triggered a marked increase in CRE-mediated gene expression in the SCN. Reporter gene experiments showed that activation of the p44/42 mitogen-activated protein kinase signaling cascade is required for Ca 2؉-dependent stimulation of CRE-mediated transcription in the SCN. These findings reveal the CREB/CRE transcriptional pathway to be circadian-regulated within the SCN, and raise the possibility that this pathway provides signaling information essential for normal clock function.
Memory retrieval is a dynamic aspect of memory formation that can be studied separately from othe... more Memory retrieval is a dynamic aspect of memory formation that can be studied separately from other stages of memory processing. Although several signal transduction pathways including ERK/MAP kinase have been implicated in memory retrieval, the underlying signaling events are poorly defined. Here we report that re-exposure of mice to context after contextual training stimulates the activity of phosphatidylinositol 3 kinase (PI3K) in the hippocampus. Inhibition of PI3K activity in the hippocampus in vivo blocked contextual memory retrieval and extinction. Inhibitors of PI3K signaling also blocked increases in ERK/MAP kinase activity associated with memory retrieval. This suggests that PI3K activation in the hippocampus is critical for memory retrieval and is required for activation of ERK/MAP kinase during retrieval. Understanding the means by which the nervous system can store and manage vast amounts of information remains a central goal in neuroscience. Memory can be experimentally divided into several stages, including acquisition, consolidation, retrieval, reconsolidation and extinction 1-3. An initial memory formed during acquisition is unstable and is subsequently consolidated by a process that requires de novo gene expression and protein synthesis 4,5. During contextual memory formation, an animal that is shocked in context develops a strong memory for context manifested by increased freezing when returned to the training context. When a trained animal is brought back into the training context without a shock, the memory is retrieved, which results in either reconsolidation or extinction depending upon the strength and age of the memory as well as the duration of retrieval 3,6,7. Although the different stages of memory formation may share some common mechanisms, it has become increasingly evident that the underlying molecular mechanisms differ. Memory retrieval is an essential component of memory processing; without retrieval, memories are not expressed. Although there is controversy concerning the molecular events required for memory retrieval 1-3 , several regulatory pathways including ERK/MAP kinase (ERK/MAPK) and cAMP signaling are strongly implicated 8,9. Furthermore, contextual memory retrieval evokes a marked induction of c-Fos and JunB in pyramidal CA1 neurons of the dorsal hippocampus 10. In contrast to memory acquisition and consolidation, memory retrieval does not, however, depend on activation of NMDA receptors or CaM kinase II (refs. 8,11). Another activity-stimulated signaling pathway implicated in memory formation and synaptic plasticity is PI3K. PI3K signaling is required for long-term potentiation (LTP) in different areas of brain, including the Schaffer collateral/commissural fiber CA1 synapses 12,13 , dentate gyrus 14 and amygdala 15. In addition, injection of PI3K inhibi
of them are due to defects in the cAMP pathway: amnesiac, a putative adenylyl cyclase-activating ... more of them are due to defects in the cAMP pathway: amnesiac, a putative adenylyl cyclase-activating peptide (Feany and Quinn, 1995); dunce, a cAMP phosphodiesterase (Chen et al., 1986); rutabaga, a Ca 2ϩ-stimulated adenylyl cyclase (Livingston et al., 1984); and DCO, a
This report was funded by the Bonneville Power Administration (BPA), U.S. Department of Energy, a... more This report was funded by the Bonneville Power Administration (BPA), U.S. Department of Energy, as part of BPA's program to protect, mitigate, and enhance fish and wildlife affected by the development and operation of hydroelectric facilities on the Columbia River and its tributaries. The views in this report are the author's and do not necessarily represent the views of BPA.
ment binding protein (dCREB2-b) in Drosophila is reported to disrupt LTM (Yin et al., 1994) where... more ment binding protein (dCREB2-b) in Drosophila is reported to disrupt LTM (Yin et al., 1994) whereas expression of an activator isoform of CREB (dCREB2-a) enhances LTM (Yin et al., 1995). Related studies in mice have also implicated cAMP
Specific subcellular targeting and spatial arrangement of signaling molecules are important for e... more Specific subcellular targeting and spatial arrangement of signaling molecules are important for efficient signal transduction. The neuro-specific type-I adenylyl cyclase (AC1) is stimulated by Ca 2+ , and plays an essential role in neurodevelopment and neuroplasticity. We generated hemagglutinin (HA)-tagged AC1 to study its subcellular localization in cultured neurons. The HA-tagged AC1 has similar enzymatic activity and regulatory properties to that of non-tagged protein. HA-AC1 targeted to both apical and basolateral domains in the epithelial Madin-Darby canine kidney (MDCK) cells, and it was found in both axons and dendrites in cultured hippocampal neurons as well as in cerebellar granule neurons. Interestingly, AC1 showed a distinct punctate form of immunostaining in MDCK cells and transfected neurons, suggesting it targets to specific subcellular domains. By immunostaining with different synaptic markers, we found that AC1 puncta were located at the excitatory synapses in cerebellar granule neurons. Our data provide a possible cellular mechanism for the physiological role of AC1 in neuroplasticity.
The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in ... more The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in intact coho salmon follicles and isolated follicular layers at various stages of oocyte maturation, from late vitellogenesis until the completion of germinal vesicle breakdown (GVBD). In granulosa layers from all stages, LH, but not FSH, stimulated 17␣,20-dihydroxy-4-pregnen-3-one (17,20-P) production. In theca-interstitial layers from all stages, FSH and LH stimulated steroid production, LH being more potent than FSH. The basal steroid output of intact follicles was significantly lower than that of isolated follicular layers, and their response to FSH and LH also differed. First, the intact follicles produced 17␣-hydroxyprogesterone in response to FSH during the central germinal vesicle stage while thecainterstitial layers did not. Second, estradiol-17 production was not inhibited by LH during final oocyte maturation in intact follicles, as observed for granulosa layers. Our results indicate that LH is the determining factor regulating the production of the maturation-inducing steroid, 17,20-P, and the induction of GVBD in the salmonid ovary. In summary, we have provided evidence for maturation-associated changes in the effects of FSH and LH in the salmonid ovary, which further supports the hypothesis that FSH and LH have distinct functions in the teleost ovary.
A program of stringently-regulated gene expression is thought to be a fundamental component of th... more A program of stringently-regulated gene expression is thought to be a fundamental component of the circadian clock. Although recent work has implicated a role for E-box-dependent transcription in circadian rhythmicity, the contribution of other enhancer elements has yet to be assessed. Here, we report that cells of the suprachiasmatic nuclei (SCN) exhibit a prominent circadian oscillation in cAMP response element (CRE)-mediated gene expression. Maximal reporter gene expression occurred from late-subjective night to mid-subjective day. Cycling of CRE-dependent transcription was not observed in other brain regions, including the supraoptic nucleus and piriform cortex. Levels of the phospho-active form of the transcription factor CREB (P-CREB) varied as a function of circadian time. Peak P-CREB levels occurred during the mid-to late-subjective night. Furthermore, photic stimulation during the subjective night, but not during the subjective day, triggered a marked increase in CRE-mediated gene expression in the SCN. Reporter gene experiments showed that activation of the p44/42 mitogen-activated protein kinase signaling cascade is required for Ca 2؉-dependent stimulation of CRE-mediated transcription in the SCN. These findings reveal the CREB/CRE transcriptional pathway to be circadian-regulated within the SCN, and raise the possibility that this pathway provides signaling information essential for normal clock function.
The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in ... more The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in intact coho salmon follicles and isolated follicular layers at various stages of oocyte maturation, from late vitellogenesis until the completion of germinal vesicle breakdown (GVBD). In granulosa layers from all stages, LH, but not FSH, stimulated 17␣,20-dihydroxy-4-pregnen-3-one (17,20-P) production. In theca-interstitial layers from all stages, FSH and LH stimulated steroid production, LH being more potent than FSH. The basal steroid output of intact follicles was significantly lower than that of isolated follicular layers, and their response to FSH and LH also differed. First, the intact follicles produced 17␣-hydroxyprogesterone in response to FSH during the central germinal vesicle stage while thecainterstitial layers did not. Second, estradiol-17 production was not inhibited by LH during final oocyte maturation in intact follicles, as observed for granulosa layers. Our results indicate that LH is the determining factor regulating the production of the maturation-inducing steroid, 17,20-P, and the induction of GVBD in the salmonid ovary. In summary, we have provided evidence for maturation-associated changes in the effects of FSH and LH in the salmonid ovary, which further supports the hypothesis that FSH and LH have distinct functions in the teleost ovary.
Specific subcellular targeting and spatial arrangement of signaling molecules are important for e... more Specific subcellular targeting and spatial arrangement of signaling molecules are important for efficient signal transduction. The neuro-specific type-I adenylyl cyclase (AC1) is stimulated by Ca 2+ , and plays an essential role in neurodevelopment and neuroplasticity. We generated hemagglutinin (HA)-tagged AC1 to study its subcellular localization in cultured neurons. The HA-tagged AC1 has similar enzymatic activity and regulatory properties to that of non-tagged protein. HA-AC1 targeted to both apical and basolateral domains in the epithelial Madin-Darby canine kidney (MDCK) cells, and it was found in both axons and dendrites in cultured hippocampal neurons as well as in cerebellar granule neurons. Interestingly, AC1 showed a distinct punctate form of immunostaining in MDCK cells and transfected neurons, suggesting it targets to specific subcellular domains. By immunostaining with different synaptic markers, we found that AC1 puncta were located at the excitatory synapses in cerebellar granule neurons. Our data provide a possible cellular mechanism for the physiological role of AC1 in neuroplasticity.
A program of stringently-regulated gene expression is thought to be a fundamental component of th... more A program of stringently-regulated gene expression is thought to be a fundamental component of the circadian clock. Although recent work has implicated a role for E-box-dependent transcription in circadian rhythmicity, the contribution of other enhancer elements has yet to be assessed. Here, we report that cells of the suprachiasmatic nuclei (SCN) exhibit a prominent circadian oscillation in cAMP response element (CRE)-mediated gene expression. Maximal reporter gene expression occurred from late-subjective night to mid-subjective day. Cycling of CRE-dependent transcription was not observed in other brain regions, including the supraoptic nucleus and piriform cortex. Levels of the phospho-active form of the transcription factor CREB (P-CREB) varied as a function of circadian time. Peak P-CREB levels occurred during the mid-to late-subjective night. Furthermore, photic stimulation during the subjective night, but not during the subjective day, triggered a marked increase in CRE-mediated gene expression in the SCN. Reporter gene experiments showed that activation of the p44/42 mitogen-activated protein kinase signaling cascade is required for Ca 2؉-dependent stimulation of CRE-mediated transcription in the SCN. These findings reveal the CREB/CRE transcriptional pathway to be circadian-regulated within the SCN, and raise the possibility that this pathway provides signaling information essential for normal clock function.
Memory retrieval is a dynamic aspect of memory formation that can be studied separately from othe... more Memory retrieval is a dynamic aspect of memory formation that can be studied separately from other stages of memory processing. Although several signal transduction pathways including ERK/MAP kinase have been implicated in memory retrieval, the underlying signaling events are poorly defined. Here we report that re-exposure of mice to context after contextual training stimulates the activity of phosphatidylinositol 3 kinase (PI3K) in the hippocampus. Inhibition of PI3K activity in the hippocampus in vivo blocked contextual memory retrieval and extinction. Inhibitors of PI3K signaling also blocked increases in ERK/MAP kinase activity associated with memory retrieval. This suggests that PI3K activation in the hippocampus is critical for memory retrieval and is required for activation of ERK/MAP kinase during retrieval. Understanding the means by which the nervous system can store and manage vast amounts of information remains a central goal in neuroscience. Memory can be experimentally divided into several stages, including acquisition, consolidation, retrieval, reconsolidation and extinction 1-3. An initial memory formed during acquisition is unstable and is subsequently consolidated by a process that requires de novo gene expression and protein synthesis 4,5. During contextual memory formation, an animal that is shocked in context develops a strong memory for context manifested by increased freezing when returned to the training context. When a trained animal is brought back into the training context without a shock, the memory is retrieved, which results in either reconsolidation or extinction depending upon the strength and age of the memory as well as the duration of retrieval 3,6,7. Although the different stages of memory formation may share some common mechanisms, it has become increasingly evident that the underlying molecular mechanisms differ. Memory retrieval is an essential component of memory processing; without retrieval, memories are not expressed. Although there is controversy concerning the molecular events required for memory retrieval 1-3 , several regulatory pathways including ERK/MAP kinase (ERK/MAPK) and cAMP signaling are strongly implicated 8,9. Furthermore, contextual memory retrieval evokes a marked induction of c-Fos and JunB in pyramidal CA1 neurons of the dorsal hippocampus 10. In contrast to memory acquisition and consolidation, memory retrieval does not, however, depend on activation of NMDA receptors or CaM kinase II (refs. 8,11). Another activity-stimulated signaling pathway implicated in memory formation and synaptic plasticity is PI3K. PI3K signaling is required for long-term potentiation (LTP) in different areas of brain, including the Schaffer collateral/commissural fiber CA1 synapses 12,13 , dentate gyrus 14 and amygdala 15. In addition, injection of PI3K inhibi
of them are due to defects in the cAMP pathway: amnesiac, a putative adenylyl cyclase-activating ... more of them are due to defects in the cAMP pathway: amnesiac, a putative adenylyl cyclase-activating peptide (Feany and Quinn, 1995); dunce, a cAMP phosphodiesterase (Chen et al., 1986); rutabaga, a Ca 2ϩ-stimulated adenylyl cyclase (Livingston et al., 1984); and DCO, a
This report was funded by the Bonneville Power Administration (BPA), U.S. Department of Energy, a... more This report was funded by the Bonneville Power Administration (BPA), U.S. Department of Energy, as part of BPA's program to protect, mitigate, and enhance fish and wildlife affected by the development and operation of hydroelectric facilities on the Columbia River and its tributaries. The views in this report are the author's and do not necessarily represent the views of BPA.
ment binding protein (dCREB2-b) in Drosophila is reported to disrupt LTM (Yin et al., 1994) where... more ment binding protein (dCREB2-b) in Drosophila is reported to disrupt LTM (Yin et al., 1994) whereas expression of an activator isoform of CREB (dCREB2-a) enhances LTM (Yin et al., 1995). Related studies in mice have also implicated cAMP
Specific subcellular targeting and spatial arrangement of signaling molecules are important for e... more Specific subcellular targeting and spatial arrangement of signaling molecules are important for efficient signal transduction. The neuro-specific type-I adenylyl cyclase (AC1) is stimulated by Ca 2+ , and plays an essential role in neurodevelopment and neuroplasticity. We generated hemagglutinin (HA)-tagged AC1 to study its subcellular localization in cultured neurons. The HA-tagged AC1 has similar enzymatic activity and regulatory properties to that of non-tagged protein. HA-AC1 targeted to both apical and basolateral domains in the epithelial Madin-Darby canine kidney (MDCK) cells, and it was found in both axons and dendrites in cultured hippocampal neurons as well as in cerebellar granule neurons. Interestingly, AC1 showed a distinct punctate form of immunostaining in MDCK cells and transfected neurons, suggesting it targets to specific subcellular domains. By immunostaining with different synaptic markers, we found that AC1 puncta were located at the excitatory synapses in cerebellar granule neurons. Our data provide a possible cellular mechanism for the physiological role of AC1 in neuroplasticity.
The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in ... more The regulation of ovarian steroidogenesis in vitro by coho salmon FSH and LH was investigated in intact coho salmon follicles and isolated follicular layers at various stages of oocyte maturation, from late vitellogenesis until the completion of germinal vesicle breakdown (GVBD). In granulosa layers from all stages, LH, but not FSH, stimulated 17␣,20-dihydroxy-4-pregnen-3-one (17,20-P) production. In theca-interstitial layers from all stages, FSH and LH stimulated steroid production, LH being more potent than FSH. The basal steroid output of intact follicles was significantly lower than that of isolated follicular layers, and their response to FSH and LH also differed. First, the intact follicles produced 17␣-hydroxyprogesterone in response to FSH during the central germinal vesicle stage while thecainterstitial layers did not. Second, estradiol-17 production was not inhibited by LH during final oocyte maturation in intact follicles, as observed for granulosa layers. Our results indicate that LH is the determining factor regulating the production of the maturation-inducing steroid, 17,20-P, and the induction of GVBD in the salmonid ovary. In summary, we have provided evidence for maturation-associated changes in the effects of FSH and LH in the salmonid ovary, which further supports the hypothesis that FSH and LH have distinct functions in the teleost ovary.
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