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Biomedical subjects

C L Williams

Publications and source records attributed to C L Williams.

At least 37 records · Page 2Linked to original sources
PubMed2000

P2Y2 purinergic and M3 muscarinic acetylcholine receptors activate different phospholipase C-beta isoforms that are uniquely susceptible to protein kinase C-dependent phosphorylation and inactivation.

Activation of phospholipase C-beta (PLC-beta) by G protein-coupled receptors typically results in rapid but transient second messenger generation. Although PLC-beta deactivation may contribute to the transient nature of this response, the mechanisms governing PLC-beta deactivation are poorly characterized. We investigated the involvement of protein kinase C (PKC) in the termination of PLC-beta activation induced by endogenous P2Y(2) purinergic receptors and transfected M(3) muscarinic acetylcholine receptors (mAChR) in Chinese hamster ovary cells. Activation of P2Y(2) receptors causes Galpha(q/11) to associate with PLC-beta3, whereas M(3) mAChR activation causes Galpha(q/11) to associate with both PLC-beta1 and PLC-beta3 in these cells. Phosphorylation of PLC-beta3, but not PLC-beta1, is induced by activating either P2Y(2) receptors or M(3) mAChR. We demonstrate that PKC rather than protein kinase A mediates the G protein-coupled receptor-induced phosphorylation of PLC-beta3. The PKC-mediated phosphorylation of PLC-beta3 diminishes the interaction of Galpha(q/11) with PLC-beta3, thereby contributing to the termination PLC-beta3 activity. These findings indicate that the distinct temporal profiles of PLC activation by P2Y(2) receptors and mAChR may arise from the differential activation of PLC-beta1 and PLC-beta3 by the receptors, coupled with a selective PKC-mediated negative feedback mechanism that targets PLC-beta3 but not PLC-beta1.

Animals↗
PubMed2000

Prenatal choline exposure alters hippocampal responsiveness to cholinergic stimulation in adulthood.

Manipulation of dietary choline levels during gestation results in enduring neurobehavioral changes in offspring that last into adulthood. Alterations of hippocampal function and memory are among the most striking changes. Depending upon the measures assessed, prenatal choline supplementation tends to promote excitatory synaptic efficacy in hippocampal circuits while prenatal choline deficiency diminishes it. However, the mechanisms underlying these changes remain unclear. Transverse hippocampal slices were prepared from adult offspring of dams fed choline supplemented, choline deficient, or control diets. We assessed paired-pulse inhibition, and excitatory synaptic responsiveness before and after activation of cholinergic receptors with Carbachol. Prenatally choline deficient animals yielded significantly fewer electrophysiological viable hippocampal slices than did animals from either of the other two treatment groups. Among the slices tested, there were no differences in paired pulse inhibition between the treatment groups. However, transient cholinergic activation resulted in a prolonged enhancement of the amplitude of the population EPSP (pEPSP) response in slices from prenatally choline supplemented animals. These results suggest that GABA receptor-mediated inhibition remains intact after prenatal choline manipulations, and that enhancement of the excitatory responsiveness of hippocampal circuits in slices from prenatally choline supplemented rats may be related in part to an increase in cholinergic tone within the CA1 circuit.

Animals↗
PubMed2000

Small cell lung carcinoma exhibits greater phospholipase C-beta1 expression and edelfosine resistance compared with non-small cell lung carcinoma.

Aberrant signal transduction pathways involved in the development of metastatic disease are poorly defined in both small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC). Neuropeptide-driven positive feedback loops stimulating cell proliferation are characteristic of SCLC. The activation of phospholipase C (PLC)-beta1 is an early and common response to stimulation of G protein-coupled receptors by these neuroendocrine growth factors. The importance of PLC-beta in neuropeptide signaling prompted us to compare PLC-beta isoform expression and activity in four independent SCLC cell lines and four independent NSCLC cell lines. We found that PLC-beta1 is more highly expressed in SCLC than in NSCLC, as indicated by Western blotting of cell lysates. All SCLC lines studied express PLC-beta1; only one of the NSCLC lines investigated showed detectable levels of the enzyme. NSCLC lines are significantly more sensitive to the antiproliferative effects of ET-18-OCH3 (edelfosine) compared with the SCLC lines, as indicated by [3H]thymidine uptake. The only SCLC cell line (NCI-H345) that is as sensitive as the NSCLC cell lines to ET-18-OCH3 also expresses uniquely low levels of PLC-beta1. The participation of PLC-beta1 in signaling by SCLC growth factor receptors is indicated by our finding that PLC-beta1 (but not PLC-beta3) coimnunoprecipitates with G(alpha)q/11 upon activation of neurotensin receptors; this association is inhibited by ET-18-OCH3. Ca2+ mobilization mediated by neurotensin receptors is also inhibited by ET-18-OCH3. The binding of GTPgammaS to G(alpha)q/11 upon treatment of SCLC cells with neurotensin is not inhibited by ET-18-OCH3. These findings indicate that ET-18-OCH3 does not interfere with G(alpha)q/11 activation but rather inhibits the association of G(alpha)q/11 with PLC-beta1. Our data suggest that PLC-beta is an important mediator of both SCLC and NSCLC proliferation. Differences in PLC-beta1 expression may be exploitable in the development of effective diagnostic and therapeutic tools.

Antineoplastic Agents↗
PubMed2000

Unique in vivo associations with SmgGDS and RhoGDI and different guanine nucleotide exchange activities exhibited by RhoA, dominant negative RhoA(Asn-19), and activated RhoA(Val-14).

We compared the in vivo characteristics of hemagglutinin (HA)-tagged RhoA, dominant negative RhoA(Asn-19), and activated RhoA(Val-14) stably expressed in Chinese hamster ovary (CHO) cells. Proteins co-precipitating with these HA-tagged GTPases were identified by peptide sequencing or by Western blotting. Dominant negative RhoA(Asn-19) co-precipitates with the guanine nucleotide exchange factor (GEF) SmgGDS but does not detectably interact with other expressed GEFs, such as Ost or Dbl. SmgGDS co-precipitates minimally with wild-type RhoA and does not detectably associate with RhoA(Val-14). The guanine nucleotide dissociation inhibitor RhoGDI co-precipitates with RhoA, and to a lesser extent with RhoA(Val-14), but does not detectably co-precipitate with RhoA(Asn-19). Wild-type RhoA is predominantly in the [(32)P]GDP-bound form, RhoA(Val-14) is predominantly in the [(32)P]GTP-bound form, and negligible levels of [(32)P]GDP or [(32)P]GTP are bound to RhoA(Asn-19) in (32)P-labeled cells. Immunofluorescence analyses indicate that HA-RhoA(Asn-19) is excluded from the nucleus and cell junctions. Microinjection of SmgGDS cDNA into CHO cells stably expressing HA-RhoA causes HA-RhoA to be excluded from the nucleus and cell junctions, similar to the distribution of RhoA(Asn-19). Our findings indicate that the expression of RhoA(Asn-19) may specifically inhibit signaling pathways that rely upon the SmgGDS-dependent activation of RhoA.

Amino Acid Sequence↗
PubMed2000

Posttraining inactivation of excitatory afferent input to the locus coeruleus impairs retention in an inhibitory avoidance learning task.

These experiments examined whether the nucleus paragigantocellularis (PGi) contributes to memory storage processing via its ascending excitatory influence on locus coeruleus (LC) neuronal activity. Activation of the LC leads to memory enhancement and also results in a widespread release of norepinephrine in target structures, such as the amygdala and hippocampus. Infusion of norepinephrine into either structure also improves memory for several types of learned responses. Thus, the capacity for norepinephrine to modulate memory within limbic structures may be contingent upon the functional connections between PGi and the LC. To examine this hypothesis, male Sprague-Dawley rats were implanted with cannula aimed above PGi (Experiments 1 and 2) or 1.5 mm dorsal or medial to PGi (Experiment 3). Immediately following inhibitory avoidance training (0.45 mA, 0. 5 s), phosphate-buffered saline, lidocaine (Experiment 1), or 12.5 or 25 nmol/0.5 microl of the GABA agonist muscimol (Experiment 2) was infused into PGi. On a retention test given 48 h later, the latency to reenter the footshock compartment was significantly shorter for subjects given either lidocaine or 12.5 or 25.0 nmol of muscimol compared to controls. In Experiment 3, infusion of lidocaine or muscimol into areas 1.5 mm dorsal or medial to PGi did not significantly alter retention, indicating that the memory impairment observed in Experiments 1 and 2 was site specific and not due to the spread of drug to cell groups surrounding PGi. These findings suggest that PGi may serve a vital function in relaying biologically relevant information to forebrain structures involved in memory via its excitatory influence on the LC.

Anesthetics, Local↗
PubMed2000

Noradrenergic receptor blockade of the NTS attenuates the mnemonic effects of epinephrine in an appetitive light-dark discrimination learning task.

These experiments examined the contribution of noradrenergic neurons in the nucleus of the solitary tract (NTS) in mediating the memory-facilitating effects of epinephrine. In Experiment 1, saline or 0.05 or 0.1 mg/kg of epinephrine was given intraperitoneally (ip) to rats after the second day of training in a light-dark Y-maze discrimination task. On a 20-trial retention test given 2 and 7 days later, the 0.1 mg/kg epinephrine group made significantly more correct responses than controls and required fewer trials to reach criterion. In Experiment 2, phosphate-buffered saline or the noradrenergic antagonist dl-propranolol (0.3 or 1.0 microg/0.5 microl) was infused into the NTS prior to an ip injection of saline or 0.1 mg/kg of epinephrine. The memory-enhancing effects of epinephrine were attenuated by the infusion of 0.3 microg/0.5 microl of dl-propranolol into the NTS. These findings indicate an involvement of NTS noradrenergic neurons in mediating the effects of peripheral epinephrine on memory storage processes.

Animals↗
PubMed2000

Adrenergic activation of the nucleus tractus solitarius potentiates amygdala norepinephrine release and enhances retention performance in emotionally arousing and spatial memory tasks.

It is well documented that noradrenergic systems in the amygdala modulate memory formation, however, less research has examined how sources of limbic norepinephrine contribute to this process. The amygdala receives a dense supply of norepinephrine from neurons in the nucleus of the solitary tract (NTS). The present experiments examined whether adrenergic activation of these NTS neurons affects memory in learning tasks that are sensitive to amygdala norepinephrine release. Separate groups of male Sprague-Dawley rats were trained in either an emotionally arousing or spatial memory task. They then received vehicle or the adrenergic agonist epinephrine (50, 125, or 250 ng/0.5 microl) into the NTS. Rats given the 125 ng dose had significantly longer retention latencies on a 48 h inhibitory avoidance retention test and made a significantly higher percentage of correct responses on an 18 h delayed radial maze retention test. A third experiment using in vivo microdialysis and high performance liquid chromatography (HPLC) demonstrated that intra-NTS infusion of a memory-enhancing dose of epinephrine potentiated amygdala norepinephrine release. Collectively, these results suggest that stimulation of the NTS contributes to memory processing by influencing noradrenergic systems in the amygdala.

Adrenergic Agonists↗
PubMed2000

Do friendships change behaviors, or do behaviors change friendships? Examining paths of influence in young adolescents' alcohol use.

PURPOSE: This study examined support for models of peer influence, which postulates that young adolescents whose friends use alcohol will also engage in that behavior, and of peer selection, whereby young adolescents seek out friends whose drinking behavior is similar to their own. METHODS: Data for this study are from 1804 adolescents participating in Project Northland, a school- and community-based alcohol use prevention trial. Using latent variable structural equation modeling, a series of models examined directions of influence between participant alcohol use and friend drug use over three points in Grades 7, 8, and 9. RESULTS: Findings indicated that higher levels of friends' drug use led to increased participant alcohol use. The reverse-order relationship (i.e., greater participant involvement in alcohol leading to more drug use among friends) was not supported by these data. Finally, best-fitting models supported the notion that both participants' alcohol use and the alcohol and other drug use of friends were highly stable over time. CONCLUSIONS: Similarity in drinking behavior among adolescent friends may be more related to processes of peer influence than to processes of peer selection. Findings support the utility of alcohol use prevention programs that equip younger teens with skills to resist peer influences to use alcohol.

Adolescent↗
PubMed2000

Glutamatergic influences on the nucleus paragigantocellularis: contribution to performance in avoidance and spatial memory tasks.

Stimulation of the locus coeruleus (LC) and the subsequent release of norepinephrine contribute to memory consolidation processes. Excitatory input to the LC is derived primarily from neurons in the nucleus paragigantocellularis (PGi). The authors examined the effects of activating the pathway between PGi and the LC on memory. Rats received vehicle or the excitatory amino acid glutamate (25, 50, or 100 nmol/0.5 microl) into PGi after training in an inhibitory avoidance (IA) or delayed matching-to-sample (DMS) task. Rats given the 100-nmol dose had significantly longer retention latencies on a 48-hr IA retention test. Rats treated with the 50- or 100-nmol dose made significantly more correct responses than controls on an 18-hr DMS retention test. Results suggest that encoding and storage of memory for emotional and spatial events may be enhanced by activation of neuronal circuits afferent to the LC.

Afferent Pathways↗
PubMed2000

The effects of noradrenergic activation of the nucleus tractus solitarius on memory and in potentiating norepinephrine release in the amygdala.

Although it is known that norepinephrine (NE) modulates memory by acting on limbic areas, few studies describe how structures supplying NE to the limbic system, such as the nucleus tractus solitarius (NTS) contribute to this process. The present study examined the effects on memory of activating the NE pathway between the NTS and the amygdala (AMYG). Rats received buffer or the beta-noradrenergic agonist clenbuterol (CLN; 10, 50, or 100 ng/0.5 microl) into the NTS after footshock training in a Y-maze discrimination task. Infusion of 100 ng CLN significantly improved memory when retention was tested in the absence or presence of cues associated with the footshock. Experiment 2 used in vivo microdialysis to determine whether the mnemonic effects of CLN are mediated by influencing NE output in the AMYG. Subjects were given an intra-NTS infusion of CLN or phosphate buffered saline, footshock (0.8 mA, 1 s), and injected with epinephrine (EPI; 0.3 mg/kg ip) or saline. CLN or EPI injection produced a significant increase in NE sampled from the AMYG. These findings indicate that activation of NTS neurons that project to and release NE in the AMYG modulates memory storage processing.

Amygdala↗
PubMed2000

Comparison of an enzyme immunoassay for the detection of Helicobacter pylori antigens in the faeces with the urea breath test.

BACKGROUND: Current diagnostic tests for Helicobacter pylori are invasive (endoscopy) or indirect (urea breath test, serology). AIMS: To evaluate a new enzyme immunoassay (EIA) which detects H pylori antigens in faeces, by comparing its sensitivity and specificity in children with the (13)C urea breath test (UBT). METHODS: A total of 119 children underwent a UBT and provided a faecal sample for antigen testing within seven days. After an overnight fast each child provided a pretest breath sample, and samples at 30 and 40 minutes after ingestion of 100 mg (13)C labelled urea. (13)C enrichment of breath was measured by isotope ratio mass spectrometry. Faeces were stored at -70 degrees C until antigen testing, using the EIA. Samples were read spectrophotometrically at 450 nm and results were interpreted using recommended cut offs of optical density <0.14 as negative, >/=0.16 as positive, with >/=0.14 and <0. 16 representing equivocal results. Sensitivity and specificity were calculated using the manufacturer's cut off compared with UBT. RESULTS: Sensitivity and specificity were 88% and 82%, respectively. Negative and positive predictive values were 97% and 58%. CONCLUSIONS: The EIA is an alternative, non-invasive, and easy to use method for the detection of H pylori in children. Its high negative predictive value suggests a role in screening out uninfected children.

Adolescent↗
PubMed2000

Project Northland high school interventions: community action to reduce adolescent alcohol use.

Project Northland is a randomized community trial initially implemented in 24 school districts and communities in northeastern Minnesota, with goals of delaying onset and reducing adolescent alcohol use using community-wide, multiyear, multiple interventions. The study targets the Class of 1998 from the 6th to 12th grades (1991-1998). The early adolescent phase of Project Northland has been completed, and reductions in the prevalence of alcohol use at the end of 8th grade were achieved. Phase II of Project Northland, targeting 11th- and 12th-grade students, uses five major strategies: (1) direct action community organizing methods to encourage citizens to reduce underage access to alcohol, (2) youth development involving high school students in youth action teams, (3) print media to support community organizing and youth action initiatives and communicate healthy norms about underage drinking (e.g., providing alcohol to minors is unacceptable), (4) parent education and involvement, and (5) a classroom-based curriculum for 11th-grade students. This article describes the background, design, implementation, and process measures of the intervention strategies for Phase II of Project Northland.

Adolescent↗
PubMed1999

Choline availability to the developing rat fetus alters adult hippocampal long-term potentiation.

Supplementation with choline during pregnancy in rats causes a long-lasting improvement of visuospatial memory of the offspring. To determine if the behavioral effects of choline are related to physiological changes in hippocampus, the effect of perinatal choline supplementation or deficiency on long-term potentiation (LTP) was examined in hippocampal slices of 6-8 and 12-14 month old rats born to dams consuming a control, choline-supplemented, or a choline-free diet during pregnancy. Stimulating and recording electrodes were placed in stratum radiatum of area CA1 to record extracellular population excitatory postsynaptic potentials (pEPSPs). To induce LTP, a theta-like stimulus train was generated. The amplitude of the stimulus pulses was set at either 10% or 50% of the stimulus intensity which had induced the maximal pEPSP slope on the input/output curve. We found that at both ages, a significantly smaller percentage of slices from perinatally choline-deficient rats displayed LTP after 10% stimulus intensity (compared with control and choline-supplemented rats), and a significantly larger percentage of slices from choline-supplemented rats displayed LTP at 50% stimulus intensity (compared with control and choline-deficient rats). Results reveal that alterations in the availability of dietary choline during discrete periods of development lead to changes in hippocampal electrophysiology that last well into adulthood. These changes in LTP threshold may underlie the observed enhancement of visuospatial memory seen after prenatal choline supplementation and point to the importance of choline intake during pregnancy for development of brain and memory function.

Aging↗
PubMed1999

Choline supplementation during prenatal development reduces proactive interference in spatial memory.

Previous research has demonstrated that increasing dietary choline during early development can have long-lasting effects on cholinergic (Ch) function that are correlated with improvement of spatial memory ability in rats. The present study is designed to further our understanding of these organizational changes in brain and behavior by examining the effects of spaced vs. massed trials. A third of the rats (n=10) were supplemented with choline chloride prenatally by adding it to the drinking water of their dams. Another third were made deficient of choline during early development by removing choline from the dams diet. The remaining rats served as untreated controls. Postnatally, the offspring were maintained on a choline-sufficient diet and at 120 days of age they began 12-arm radial maze training. The maze data revealed two major effects of early choline availability: (1) Both choline-supplemented and choline-deficient rats performed more accurately than control littermates when trials were spaced. These differences in spatial ability did not appear to be a function of differential response or cue-use strategies. (2) Choline-supplemented rats showed little proactive interference when trials were massed; whereas control rats demonstrated moderate levels and choline-deficient rats exhibited high levels of proactive interference as a function of massed trials. These data suggest that the behavioral consequences of early dietary availability of choline may involve the modification of the discriminative abilities used to attend to stimuli that demarcate the end of one trial and the start of another as well as the capacity for remembering the locations that have been visited during a trial.

Animals↗
PubMed1999

M3 muscarinic acetylcholine receptors regulate cytoplasmic myosin by a process involving RhoA and requiring conventional protein kinase C isoforms.

Although muscarinic acetylcholine receptors (mAChR) regulate the activity of smooth muscle myosin, the effects of mAChR activation on cytoplasmic myosin have not been characterized. We found that activation of transfected human M3 mAChR induces the phosphorylation of myosin light chains (MLC) and the formation of myosin-containing stress fibers in Chinese hamster ovary (CHO-m3) cells. Direct activation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) also induces myosin light chain phosphorylation and myosin reorganization in CHO-m3 cells. Conventional (alpha), novel (delta), and atypical (iota) PKC isoforms are activated by mAChR stimulation or PMA treatment in CHO-m3 cells, as indicated by PKC translocation or degradation. mAChR-mediated myosin reorganization is abolished by inhibiting conventional PKC isoforms with Go6976 (IC50 = 0.4 microM), calphostin C (IC50 = 2.4 microM), or chelerythrine (IC50 = 8.0 microM). Stable expression of dominant negative RhoAAsn-19 diminishes, but does not abolish, mAChR-mediated myosin reorganization in the CHO-m3 cells. Similarly, mAChR-mediated myosin reorganization is diminished, but not abolished, in CHO-m3 cells which are multi-nucleate due to inactivation of Rho with C3 exoenzyme. Expression of dominant negative RhoAAsn-19 or inactivation of RhoA with C3 exoenzyme does not affect PMA-induced myosin reorganization. These findings indicate that the PKC-mediated pathway of myosin reorganization (induced either by M3 mAChR activation or PMA treatment) can continue to operate even when RhoA activity is diminished in CHO-m3 cells. Conventional PKC isoforms and RhoA may participate in separate but parallel pathways induced by M3 mAChR activation to regulate cytoplasmic myosin. Changes in cytoplasmic myosin elicited by M3 mAChR activation may contribute to the unique ability of these receptors to regulate cell morphology, adhesion, and proliferation.

ADP Ribose Transferases↗
PubMed1999

Granule cells in aging rats are sexually dimorphic in their response to estradiol.

Normal aging comprises cognitive decline, including deterioration of memory. It has been suggested that this decline in memory is sexually dimorphic because of the cessation in gonadal steroid secretion that occurs during reproductive aging in female, but not male, mammals. We wondered whether neurons in brain regions associated with learning and memory underwent morphological changes that were dimorphic as well and whether cessation of the secretion of gonadal steroids influenced these morphological changes. To explore these questions, we deprived and restored estrogens to young and old gonadectomized females and males and studied the morphology of dentate granule cells by intracellular dye filling in a lightly fixed slice preparation. We found the following: (1) Aged female dentate granule cells deprived of gonadal steroids long-term have a paucity of dendritic spines compared with young females deprived short-term; however, aged male dentate granule cells deprived of gonadal steroids long-term have no decrease in dendritic spines compared with young males deprived short-term. (2) Aged female dentate granule cells with long-term estrogen replacement at either high or low levels still had a decline in spine density. (3) Aged female dentate granule cells with short-term estradiol replacement had spine density increased to levels normally observed in young adults, whereas aged males with short-term estradiol replacement had decreased spine density. These data suggest that the response of rat dentate granule cells to aging and estradiol is sexually dimorphic and that, in females, the responsiveness of granule cells depends on the temporal pattern of estradiol replacement.

Aging↗
PubMed1999

Activation of transfected M1 or M3 muscarinic acetylcholine receptors induces cell-cell adhesion of Chinese hamster ovary cells expressing endogenous cadherins.

Expression of endogenous cadherins by Chinese hamster ovary (CHO) cells has not been previously reported. However, we observed that CHO cells adhere to one another upon activation of transfected muscarinic acetylcholine receptors (mAChR), suggesting that the cells express endogenous cadherins. A 160-base pair RT-PCR product with 100% homology to the cytoplasmic domain of human E-cadherin was amplified from CHO cells. A second RT-PCR product amplified from these cells has 92% homology to the cytoplasmic domain of human cadherin-9 and 86% homology to the cytoplasmic domain of human cadherin-6. Western blotting indicates that CHO cells express a 165-kDa protein recognized by E-cadherin antibodies and a 120-kDa protein recognized by an antibody to the cadherin C-terminus sequence. The ability of transfected mAChR subtypes to regulate cadherin-mediated adhesion of CHO cells was tested by measuring the permeation of horseradish peroxidase across confluent CHO cell monolayers, by microscopic examination of the cells, and by aggregation assays. Cell-cell adhesion is induced within 15 min of activating transfected M1 or M3 mAChR which functionally couple to protein kinase C (PKC). In contrast, CHO cell adhesion is not affected by activating transfected M2 mAChR which functionally couple to other effectors. Activation of PKC with phorbol esters also induces cell-cell adhesion of all CHO sublines tested. Immunofluorescence assays reveal that endogenous cadherins redistribute on the plasma membrane of CHO cells following mAChR or PKC activation. Inactivation of cadherins by removal of extracellular Ca2+ abrogates adhesion induced by mAChR or PKC activation. Our demonstration that activation of only odd-numbered mAChR subtypes induces cadherin-mediated adhesion suggests that the unique responses of cells to M1 or M3 mAChR stimulation may involve cadherin activation.

Amino Acid Sequence↗