Biomedical subjects

D Kuhl

Publications and source records attributed to D Kuhl.

At least 19 recordsLinked to original sources

PubMed2001

Cerebral localization and regulation of the cell volume-sensitive serum- and glucocorticoid-dependent kinase SGK1.

The serum- and glucocorticoid-dependent kinase SGK1 is regulated by alterations of cell volume, whereby cell shrinkage increases and cell swelling decreases the transcription, expression and activity of SGK1. The kinase is expressed in all human tissues studied including the brain. The present study was performed to localize the sites of SGK1 transcription in the brain, to elucidate the influence of the hydration status on SGK1 transcription and to explore the functional significance of altered SGK1 expression. Northern blot analysis of human brain showed SGK1 to be expressed in all cerebral structures examined: amygdala, caudate nucleus, corpus callosum, hippocampus, substantia nigra, subthalamic nucleus and thalamus. In situ hybridization and immunohistochemistry in the rat revealed increased expression of SGK1 in neurons of the hippocampal area CA3 after dehydration, compared with similar slices from brains of euvolaemic rats. Additionally, several oligodendrocytes, a few microglial cells, but no astrocytes, were positive for SGK1. The abundance of SGK1 mRNA in the temporal lobe, including hippocampus, was increased by dehydration and SGK1 transcription in neuroblastoma cells was stimulated by an increase of extracellular osmolarity. Co-expression studies in Xenopus laevis oocytes revealed that SGK1 markedly increased the activity of the neuronal K+ channel Kv1.3. As activation of K+ channels modifies excitation of neuronal cells, SGK1 may participate in the regulation of neuronal excitability.

Animals↗

PubMed2001

Arg3.1/Arc mRNA induction by Ca2+ and cAMP requires protein kinase A and mitogen-activated protein kinase/extracellular regulated kinase activation.

Long-term potentiation (LTP) is a cellular model for persistent synaptic plasticity in the mammalian brain. Like several forms of memory, long-lasting LTP requires cAMP-mediated activation of protein kinase A (PKA) and is dependent on gene transcription. Consequently, activity-dependent genes such as c-fos that contain cAMP response elements (CREs) in their 5' regulatory region have been studied intensely. More recently, arg3.1/arc became of interest, because after synaptic stimulation, arg3.1/arc mRNA is rapidly induced and distributed to dendritic processes and may be locally translated there to facilitate synapse-specific modifications. However, to date nothing is known about the signaling mechanisms involved in the induction of this gene. Here we report that arg3.1/arc is robustly induced with LTP stimulation even at intensities that are not sufficient to activate c-fos expression. Unlike c-fos, the 5' regulatory region of arg3.1/arc does not contain a CRE consensus sequence and arg3.1/arc is unresponsive to cAMP in NIH3T3 and Neuro2a cells. However, in PC12 cells and primary cultures of hippocampal neurons, arg3.1/arc can be induced by cAMP and calcium. This induction requires the activity of PKA and mitogen-activated protein kinase, suggesting a neuron-specific pathway for the activation of arg3.1/arc expression.

Animals↗

PubMed2001

Two rat brain staufen isoforms differentially bind RNA.

In neurones, a limited number of mRNAs is found in dendrites, including transcripts encoding the microtubule-associated protein 2 (MAP2). Recently, we identified a cis-acting dendritic targeting element (DTE) in MAP2 mRNAs. Here we used the yeast tri-hybrid system to identify potential trans-acting RNA-binding factors of the DTE. A cDNA clone was isolated that encodes a member of a mammalian protein family that is highly homologous to the Drosophila RNA-binding protein Staufen. Mammalian Staufen appears to be expressed in most tissues and brain areas. Two distinct rat brain Staufen isoforms, rStau+I6 and rStau-I6, are encoded by alternatively spliced mRNAs. Both isoforms contain four double-stranded RNA-binding domains (dsRBD). In the larger rStau+I6 isoform, six additional amino acids are inserted in the second dsRBD. Although both isoforms interacted with the MAP2-DTE and various additional RNA fragments in an in vitro north-western assay, rStau-I6 exhibited a stronger signal of bound radioactively labelled RNAs as compared with rStau+I6. Using an antibody directed against mammalian Staufen, the protein was detected in somata and dendrites of neurones of the adult rat hippocampus and cerebral cortex. Ultrastructural studies revealed that in dendrites, rat Staufen accumulates along microtubules. Thus in neurones, rat Staufen may serve to link RNAs to the dendritic microtubular cytoskeleton and may thereby regulate their subcellular localization.

Alternative Splicing↗

PubMed2001

Are there gender differences in the use of peak flow meters?

To determine if there are gender differences in correct use of peak flow meters (PEM), third-year doctor of pharmacy students (n = 83; 52 females, 31 males) were instructed in a classroom on correct use of a PFM, including demonstrations. Students were then immediately divided into five groups, given a PFM, and assessed for three attempts in private individual sessions. Males had superior performance on the first attempt for total score (p < 0.05) and for "inhale fully" (p < 0.05). On the second attempt, the total score was not different, but males scored higher on "exhale as fast and as hard as you can" (p < 0.05). Controlled gender studies examining use of PFM in adult and pediatric patients with asthma are warranted.

Adult↗

PubMed2000

Adhesion induced expression of the serine/threonine kinase Fnk in human macrophages.

Members of the polo subfamily of protein kinases play crucial roles in cell proliferation. To study the function of this family in more detail, we isolated the cDNA of human Fnk (FGF-inducible kinase) which codes for a serine/threonine kinase of 646 aa. Despite the homology to the proliferation-associated polo-like kinase (Plk), tissue distribution of Fnk transcripts and expression kinetics differed clearly. In contrast to Plk no correlation between cell proliferation and Fnk gene expression was found. Instead high levels of Fnk mRNA were detectable in blood cells undergoing adhesion. The transition of monocytes from peripheral blood to matrix bound macrophages was accompanied by increasing levels of Fnk with time in culture. Neither treatment of monocytes with inducers of differentiation nor withdrawal of serum did influence Fnk mRNA levels significantly, suggesting that cell attachment triggers the onset of Fnk gene transcription. The idea that Fnk is part of the signalling network controlling cellular adhesion was supported by the analysis of the cytoplasmic distribution of the Fnk protein and the influence of its overexpression on the cellular architecture. Fnk as fusion protein with GFP localized at the cellular membrane in COS cells. Dysregulated Fnk gene expression disrupted the cellular f-actin network and induced a spherical morphology. Furthermore, Fnk binds to the Ca2+/integrin-binding protein Cib in two-hybrid-analyses and co-immunoprecipitation in assays. Moreover, both proteins were shown to co-localize in mammalian cells. The homology of Cib with calmodulin and with calcineurin B suggests that Cib might be a regulatory subunit of polo-like kinases.

Actins↗

PubMed2000

Odors regulate Arc expression in neuronal ensembles engaged in odor processing.

Synaptic activity is critical to developmental and plastic processes that produce long-term changes in neuronal connectivity and function. Genes expressed by neurons in an activity-dependent fashion are of particular interest since the proteins they encode may mediate neuronal plasticity. One such gene encodes the activity-regulated cytoskeleton-associated protein, Arc. The present study evaluated the effects of odor stimulation on Arc expression in rat olfactory bulb. Arc mRNA was rapidly increased in functionally linked cohorts of neurons topographically activated by odor stimuli. These included neurons surrounding individual glomeruli, mitral cells and transynaptically activated granule cells. Dendritic Arc immunoreactivity was also increased in odor-activated glomeruli. Our results suggest that odor regulation of Arc expression may contribute to activity-dependent structural changes associated with olfactory experience.

Animals↗

PubMed2000

Induction of glycerol phosphate dehydrogenase gene expression during seizure and analgesia.

Using mRNA differential display, we found that the gene for NAD(+)-dependent glycerol phosphate dehydrogenase (GPDH; EC 1.1.1.8) is induced in rat brain following seizure activity. Northern blot and in situ hybridization analysis confirmed the differential display results; they also showed, in a separate model of neuronal activation, that after thermal noxious stimulation of the hind-paws, a similar increase in GPDH mRNA occurs in the areas of somatotopic projection in the lumbar spinal cord. Surprisingly, administration of analgesic doses of morphine or the nonsteroidal antiinflammatory drugs aspirin, metamizol (dipyrone), and indomethacin also increased GPDH mRNA levels in rat spinal cord. The opioid receptor antagonist naloxone completely blocked morphine induction of GPDH but had no effect on GPDH induction by noxious heat stimulation or metamizol treatment, implicating different mechanisms of GPDH induction. Nevertheless, in all cases, induction of the GPDH gene requires adrenal steroids and new protein synthesis, as the induction was blocked in adrenalectomized rats and by cycloheximide treatment, respectively. Our results suggest that the induction of the GPDH gene upon peripheral noxious stimulation is related to the endogenous response to pain as it is mimicked by exogenously applied analgesic drugs.

Adrenalectomy↗

PubMed2000

Scatter and attenuation correction for brain SPECT using attenuation distributions inferred from a head atlas.

UNLABELLED: Sequential transmission scanning (TS)/SPECT is impractical for neurologically impaired patients who are unable to keep their heads motionless for the extended duration of the combined scans. To provide an alternative to TS, we have developed a method of inferring-attenuation distributions (IADs), from SPECT data, using a head atlas and a registration program. The validity of replacing TS with IAD was tested in 10 patients with mild dementia. METHODS: TS was conducted with each patient using a collimated 99mTc line source and fanbeam collimator; this was followed by hexamethyl propyleneamine oxime-SPECT. IAD was derived by deformably registering the brain component of a digital head atlas to a preliminary SPECT reconstruction and then applying the resulting spatial transformation to the full head atlas. SPECT data were reconstructed with scatter and attenuation correction. Relative regional cerebral blood flow was quantified in 12 threshold-guided anatomic regions of interest, with cerebellar normalization. SPECT reconstructions using IAD were compared with those using TS (which is the "gold standard") in terms of these regions of interest. RESULTS: When we compared all regions of interest across the population, the correlation between IAD-guided and TS-guided SPECT scans was 0.92 (P < 0.0001), whereas the mean absolute difference between the scans was 7.5%. On average, IAD resulted in slight underestimation of relative regional cerebral blood flow; however, this underestimation was statistically significant for only the left frontal and left central sulcus regions (P = 0.001 and 0.002, respectively). Error analysis indicated that approximately 10.0% of the total error was caused by IAD scatter correction, 36.6% was caused by IAD attenuation correction, 27.0% was caused by discrepancies in region-of-interest demarcation from quantitative errors in IAD-guided reconstructions, and 26.5% was caused by patient motion throughout the imaging procedure. CONCLUSION: SPECT reconstructions guided by IAD are sufficiently accurate to identify regional cerebral blood flow deficits of 10%, which are typical in moderate and advanced dementia.

Brain↗

PubMed1999

The polo-like protein kinases Fnk and Snk associate with a Ca(2+)- and integrin-binding protein and are regulated dynamically with synaptic plasticity.

In order to stabilize changes in synaptic strength, neurons activate a program of gene expression that results in alterations of their molecular composition and structure. Here we demonstrate that Fnk and Snk, two members of the polo family of cell cycle associated kinases, are co-opted by the brain to serve in this program. Stimuli that produce synaptic plasticity, including those that evoke long-term potentiation (LTP), dramatically increase levels of both kinase mRNAs. Induced Fnk and Snk proteins are targeted to the dendrites of activated neurons, suggesting that they mediate phosphorylation of proteins in this compartment. Moreover, a conserved C-terminal domain in these kinases is shown to interact specifically with Cib, a Ca(2+)- and integrin-binding protein. Together, these studies suggest a novel signal transduction mechanism in the stabilization of long-term synaptic plasticity.

Amino Acid Sequence↗

PubMed1999

Pim kinase expression is induced by LTP stimulation and required for the consolidation of enduring LTP.

In animals and several cellular models of synaptic plasticity, long-lasting changes in synaptic strength are dependent on gene transcription and translation. Here we demonstrate that Pim-1, a serine/threonine kinase closely related to Pim-2 and Pim-3, is induced in hippocampus in response to stimuli that evoke long-term potentiation (LTP). Mice deficient for Pim-1 show normal synaptic transmission and short-term plasticity. However, they fail to consolidate enduring LTP even though Pim-2 and Pim-3 are constitutively expressed in the hippocampus and Pim-3 expression is similarly induced by synaptic activity. Thus, expression of Pim-1 is required for LTP. Its level of expression and, consequently, its capacity to phosphorylate target proteins in dendritic and nuclear compartments of stimulated neurons might be a determining factor for the establishment of long-lasting changes in synaptic strength.

Animals↗

PubMed1999

Novelty-induced increased expression of immediate-early genes c-fos and arg 3.1 in the mouse brain.

The detection of novel stimuli is a memory-dependent process. The presented stimulus has to be compared with memory contents to judge its novelty. In addition, the novelty of stimuli activates attention-related processes that facilitate memory formation. To determine the involvement of limbic and neocortical brain structures in novelty detection, we exposed mice to a novel gustatory stimulus (0.5% saccharin) added to their drinking fluid. We then compared the novelty-induced expression of the two immediate-early genes (IEGs) c-fos and arg 3.1, with their expression in mice familiarized with the same stimulus or mice not exposed to that stimulus. Exposure to taste novelty increased expression of c-fos and arg 3.1 mRNA in the cingulate cortex and deep layers of the parietal cortex. In addition, c-fos mRNA expression was increased in the amygdala and arg 3.1 mRNA was increased in the dentate gyrus. Expression of c-fos and arg 3.1 was elevated 30 min after the exposure to novelty. For arg 3.1, a second peak of expression was found 4.5 h after presentation of the novel stimulus. Our results indicate that the amygdala, the dentate gyrus, and the cingulate and parietal cortices may be involved in novelty detection and associated cognitive events, and suggest that c-fos and arg 3.1 play distinct roles in these processes.

Animals↗

PubMed1999

Can we justify ipratropium therapy as initial management of acute exacerbations of COPD?

Although inhaled ipratropium is commonly accepted as the drug of choice for long-term management of chronic bronchitis and emphysema, little evidence is available to promote its administration in conjunction with a beta2-agonist as part of initial management of exacerbations of chronic obstructive pulmonary disease (COPD) in the acute care setting. Reasons for its widespread acceptance for acutely ill patients may include its status as a first-line agent for long-term therapy, its relative safety, and attempts to provide optimal patient care. Since inhaled ipratropium is beneficial as immediate therapy for asthma in the emergency department, some practitioners attempted to extrapolate these findings to treatment of COPD. Review of available studies reveals wide variability in methodologies and results. Although some studies reported improvement in pulmonary function tests, no clinically significant differences in patient outcomes, including shorter hospitalization, were evident. In patients who fail traditional therapies, inhaled ipratropium is reasonable. Double-blind, randomized, placebo-controlled trials in patients receiving emergency department care and in hospitalized patients that reveal shorter length of stay or other improved outcomes, are necessary to establish routine addition of inhaled ipratropium to beta2-agonists in the initial management of acute COPD.

Adrenergic beta-Agonists↗

PubMed1998

A subtractive hybridisation method for the enrichment of moderately induced sequences.

Moderately induced genes often escape detection in conventional subtraction hybridisation cloning. Here a modification of a phagemid subtraction protocol is described that overcomes this problem. The protocol uses low ratio hybridisation of driver to target sequences to allow enrichment of the sequences of interest, and back-hybridisation of the subtracted sequences with induced sequences to reduce the accumulation of false positive clones. The procedure takes advantage of the quantitative representation of cellular RNA populations in cDNA libraries, therefore, they may serve not only as renewable sources of driver and target sequences, but also as sources of population cRNAs used in northern blots and differential Southern blots.

Animals↗

PubMed1998

Dendritic localization of mRNAs.

The dendritic localization of mRNAs has been proposed to underlie the structural and functional polarity of neurons, as well as certain aspects of synaptic plasticity. Even though there is no conclusive evidence that such a localization is a physiological requirement, studies of mRNA localization in relation to function in other cell types and recent experiments on synaptic plasticity suggest that this proposal may be correct.

Animals↗

PubMed1998

Incorporating palliative care into primary care education. National Consensus Conference on Medical Education for Care Near the End of Life.

The confluence of enhanced attention to primary care and palliative care education presents educators with an opportunity to improve both (as well as patient care) through integrated teaching. Improvements in palliative care education will have benefits for dying patients and their families, but will also extend to the care of many other primary care patients, including geriatric patients and those with chronic illnesses, who make up a large proportion of the adult primary care population. In addition, caring for the dying, and teaching others to carry out this task, can be an important vehicle for personal and professional growth and development for both students and their teachers.

Curriculum↗

PubMed1996

A tri-hybrid system for the analysis and detection of RNA--protein interactions.

A modification of the two-hybrid system is described for the in vivo reconstruction of specific RNA-protein interactions. In this tri-hybrid system, the DNA binding and transcription activation domains of the yeast transcriptional activator GAL4 are brought together via the interaction of recombinant fusion proteins with a recombinant RNA. The method provides a system for studying RNA-protein interactions with the genetic advantages of the two-hybrid system. It may be used to detect specific RNA-binding proteins or target RNAs from a library of cDNAs, or to analyse the structural specificity of identified RNA-protein interactions.

Binding Sites↗

PubMed1996

A different form of long-lasting potentiation revealed in tissue plasminogen activator mutant mice.

The establishment of hippocampal long-term (LTP) requires protein and mRNA synthesis, suggesting that neuronal activity resulting in LTP initiates a cascade of gene expression. The expression of the gene for the extracellular serine protease tissue plasminogen activator (t-PA) is induced during LTP. Here we analyze long-lasting LTP (L-LTP,>4 hr)in CA1 hippocampal slices of mice homozygous for disrupted t-PA genes. Although mutant mice appear to exhibit long-term potentiation, we provide evidence that these mice are devoid of conventional homosynaptic L-LTP at the Schaffer collateral-CA1 pyramidal cell synapse. Most remarkably, t-PA-deficient mice exhibit a different form of long-lasting potentiation that is characterized by an NMDA receptor-dependent modification of GABA transmission in the CA1 region.

2-Amino-5-phosphonovalerate↗

PubMed1996

Prostatic acid phosphatase levels (enzymatic method) from completely sectioned, clinically benign, whole prostates.

Clinically benign, whole untrimmed prostates were obtained from 104 patients at autopsy, completely sectioned, and examined microscopically. The histological and gross findings of the prostate were correlated with premortem prostatic acid phosphatase levels (PAP, enzymatic method, ACA, Dupont Co.) to determine how often carcinoma of the prostate (CAP) affected PAP levels and to identify other findings within the prostate associated with elevated PAP levels. Sixty (58%) prostates did not have CAP, 34 (33%) had CAP smaller than 1 ml in volume, and 10 (10%) had CAP larger than 1 ml in volume. PAP levels were elevated (greater than 1 U/L) in 8 of 60 (13%) prostates without CAP, in 2 of the 34 (6%) prostates with CAP smaller than 1 ml, and in 1 of the 10 (10%) prostates with CAP larger than 1 ml. These differences were not statistically significant. Likewise, a statistically significant correlation between PAP levels and patient age, patient race, severe inflammation, of high grade prostatic intraepithelial neoplasia (PIN) was not found. However, there was a statistically significant correlation between PAP levels and prostate weight (p < 0.0001). This study suggest that PAP cannot distinguish between patients with clinically undetected CAP and patients without CAP. Furthermore, elevated PAP levels are often not due to metastatic CAP and additional evidence should be present, even in patients with known CAP, before an elevated PAP level is considered to be conclusive evidence of metastatic CAP.

Acid Phosphatase↗