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S Winston

Publications and source records attributed to S Winston.

At least 19 recordsLinked to original sources
PubMed2006

Electrophysiological characterization of neurons in the dorsolateral pontine rapid-eye-movement sleep induction zone of the rat: Intrinsic membrane properties and responses to carbachol and orexins.

Pharmacological, lesion and single-unit recording techniques in several animal species have identified a region of the pontine reticular formation (subcoeruleus, SubC) just ventral to the locus coeruleus as critically involved in the generation of rapid-eye-movement (REM) sleep. However, the intrinsic membrane properties and responses of SubC neurons to neurotransmitters important in REM sleep control, such as acetylcholine and orexins/hypocretins, have not previously been examined in any animal species and thus were targeted in this study. We obtained whole-cell patch-clamp recordings from visually identified SubC neurons in rat brain slices in vitro. Two groups of large neurons (mean diameter 30 and 27 mum) were tentatively identified as cholinergic (rostral SubC) and noradrenergic (caudal SubC) neurons. SubC reticular neurons (non-cholinergic, non-noradrenergic) showed a medium-sized depolarizing sag during hyperpolarizing current pulses and often had a rebound depolarization (low-threshold spike, LTS). During depolarizing current pulses they exhibited little adaptation and fired maximally at 30-90 Hz. Those SubC reticular neurons excited by carbachol (n=27) fired spontaneously at 6 Hz, often exhibited a moderately sized LTS, and varied widely in size (17-42 mum). Carbachol-inhibited SubC reticular neurons were medium-sized (15-25 mum) and constituted two groups. The larger group (n=22) was silent at rest and possessed a prominent LTS and associated one to four action potentials. The second, smaller group (n=8) had a delayed return to baseline at the offset of hyperpolarizing pulses. Orexins excited both carbachol excited and carbachol inhibited SubC reticular neurons. SubC reticular neurons had intrinsic membrane properties and responses to carbachol similar to those described for other reticular neurons but a larger number of carbachol inhibited neurons were found (>50%), the majority of which demonstrated a prominent LTS and may correspond to pontine-geniculate-occipital burst neurons. Some or all carbachol-excited neurons are presumably REM-on neurons.

Anesthetics, Local↗
PubMed2006

Differential effect of orexins (hypocretins) on serotonin release in the dorsal and median raphe nuclei of freely behaving rats.

Orexin (hypocretin)-containing neurons in the perifornical hypothalamus project to widespread regions of the brain, including the dorsal and median raphe nuclei [Peyron C, Tighe DK, van den Pol AN, de Lecea L, Heller HC, Sutcliffe JG, Kilduff TS (1998) Neurons containing hypocretin (orexin) project to multiple neuronal systems. J Neurosci 18:9996-10015; Wang QP, Koyama Y, Guan JL, Takahashi K, Kayama Y, Shioda S (2005) The orexinergic synaptic innervation of serotonin- and orexin 1-receptor-containing neurons in the dorsal raphe nucleus. Regul Pept 126:35-42]. Orexin-A or orexin-B was infused by reverse microdialysis into the dorsal raphe nucleus or median raphe nucleus of freely behaving rats, and extracellular serotonin was simultaneously collected by microdialysis and analyzed by high-performance liquid chromatography. We have found that orexin-A produced a dose-dependent increase of serotonin in the dorsal raphe nucleus, but not in the median raphe nucleus. However, orexin-B elicited a small but significant effect in both the dorsal raphe nucleus and median raphe nucleus. Orexins may have regionally selective effects on serotonin release in the CNS, implying a unique interaction between orexins and serotonin in the regulation of activities including sleep-wakefulness.

Analysis of Variance↗
PubMed2000

Vaccination of seropositive subjects with CHIRON CMV gB subunit vaccine combined with MF59 adjuvant for production of CMV immune globulin.

The safety and immunogenicity of four different regimens of CHIRON cytomegalovirus (CMV) gB subunit vaccine combined with MF59 adjuvant and administered to seropositive plasma donors were evaluated to ascertain whether vaccination of seropositive subjects would significantly increase antibody titer to gB glycoprotein. This was done to select the best vaccination regimen for generating high-titered plasma for manufacture of CMV immune globulin. No serious adverse events were attributed to this vaccine, and the vaccine was well tolerated. Only the first dose of vaccine in each regimen stimulated a four-fold or greater antibody response to gB glycoprotein and each regimen induced similar antibody titers. However, initial vaccination followed by a 1 week rest from plasmapheresis and two booster vaccinations at 8 and 24 weeks, each followed with another 1 week rest from plasmapheresis, maintained the highest geometric mean gB ELISA titer of the four regimens over the 34-week post-vaccination period. CMVIG manufactured from a pool of high titered plasma units from two of four subject groups had gB ELISA and neutralizing antibody titers nine and six times higher, respectively, compared to Cytogam, indicating that vaccination of seropositive subjects with CHIRON gB vaccine combined with MF59 adjuvant prior to harvesting plasma can enhance functional antibody in a CMVIG product.

Adjuvants, Immunologic↗
PubMed1999

REM sleep enhancement and behavioral cataplexy following orexin (hypocretin)-II receptor antisense perfusion in the pontine reticular formation.

Orexin (hypocretin)-containing neurons of the hypothalamus project to brainstem sites that are involved in the neural control of REM sleep, including the locus coeruleus, the dorsal raphe nucleus, the cholinergic zone of the mesopontine tegmentum, and the pontine reticular formation (PRF). Orexin knockout mice exhibit narcolepsy/cataplexy, and a mutant and defective gene for the orexin type II receptor is present in dogs with an inherited form of narcolepsy/cataplexy. However, the physiological systems mediating these effects have not been described. We reasoned that, since the effector neurons for the majority of REM sleep signs, including muscle atonia, were located in the PRF, this region was likely implicated in the production of these orexin-related abnormalities. To test this possibility, we used microdialysis perfusion of orexin type II receptor antisense in the PRF of rats. Ten to 24 hours after antisense perfusion, REM sleep increased two- to three-fold during both the light period (quiescent phase) and the dark period (active phase), and infrared video showed episodes of behavioral cataplexy. Moreover, preliminary data indicated no REM-related effects following perfusion with nonsense DNA, or when perfusion sites were outside the PRF. More work is needed to provide precise localization of the most effective site of orexin-induced inhibition of REM sleep phenomena.

Animals↗
PubMed1996

Pontine cholinergic neurons show Fos-like immunoreactivity associated with cholinergically induced REM sleep.

Recently, we showed c-fos expression in pontine nuclei in association with cholinergically induced REM sleep (REMc). Pontine cholinergic mechanisms have been implicated in the orchestration of the phasic and tonic events underlying REM sleep. Therefore, in the present study, we examined whether pontine cholinergic neurons demonstrate Fos-like immunoreactivity (Fos-LI) following cholinergically induced, sustained rapid-eye movement (REMc) sleep in cats. Microinjections (0.25 microliter) of vehicle (n = 2) or carbachol (n = 3; 2.0 micrograms/0.25 microliter) were made into the medial pontine reticular formation. Carbachol produced a state with all the signs of natural REM sleep, and with durations ranging from 27 to 40.1 min. Animals were killed immediately after the end of REMc. Compared to vehicle treated animals (0.9% saline), the animals with REMc showed a significantly higher number of Fos-LI cells in pontine regions implicated in REM sleep generation. More importantly, 11.2% (SEM +/- 0.83) of cholinergic neurons in the lateral dorsal tegmental (LDT) and pedunculopontine tegmental (PPT) nuclei were determined to be also Fos-LI positive. In the vehicle treated animals very few Fos-LI cells were found and none of these were found to be cholinergic. These findings indicate that during REMc a transcriptional cascade involving c-fos occurs in a subpopulation of pontine cholinergic neurons.

Acetylcholine↗
PubMed1995

Time course of phosphorylated CREB and Fos-like immunoreactivity in the hypothalamic supraoptic nucleus after salt loading.

Phosphorylation of the cAMP response element binding protein (CREB) precedes the induction of immediate early gene expression. Using antibodies that distinguish CREB from phosphorylated CREB (PCREB), we studied the appearance of PCREB-like immunoreactivity (PCREB-LI) and Fos-LI in the hypothalamic supraoptic nucleus (SON) of rats treated with hypertonic or normal saline and uninjected controls. Fifteen minutes after injection, increased numbers of PCREB-LI cells were seen in both normal and hypertonic saline-treated rats as compared with uninjected controls. Forty-five minutes after injection, levels of c-fos mRNA in the SON were elevated in hypertonic saline-treated rats as compared with normal saline-treated rats, and were minimally detectable in uninjected rats. At this time period, the hypertonic saline-treated rats showed increased number of Fos-LI cells in the SON, whereas normal saline-treated rats showed little or no Fos-LI cells. The discrepancy between levels of PCREB-LI and c-fos mRNA suggests that injection of hypertonic saline may activate additional transcriptional factors besides CREB. The lack of Fos-LI in the presence of modest increases in c-fos mRNA in normal saline-treated rats implies that levels of c-fos mRNA must exceed a threshold before increases in Fos-LI cells are detectable by immunostaining of the SON. Such a threshold might permit neuronal cells to activate diverse genes, through phosphorylation of CREB, without inducing the constellation of Fos-responsive genes.

Animals↗
PubMed1995

Effect of conjugation methodology, carrier protein, and adjuvants on the immune response to Staphylococcus aureus capsular polysaccharides.

Conjugate vaccines were prepared with S. aureus type 8 capsular polysaccharide (CP) using three carrier proteins: Pseudomonas aeruginosa exotoxin A (ETA), a non-toxic recombinant ETA (rEPA), and diphtheria toxoid (DTd). Adipic acid dihydrazide (ADH) or N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) was used as a spacer to link the CP to carrier protein. All conjugates gave a high immune response with a boost after the second immunization. Conjugates prepared with ADH gave higher antibody titers than conjugates prepared with SPDP. IgG1 was the primary subclass elicited by all conjugates regardless of the carrier protein or the conjugation method used to prepare the vaccines. The non-immunogenic CP and the conjugates were formulated with either monophosphoryl lipid A (MPL), QS21, or in Novasomes and evaluated in mice. While the adjuvants failed to improve the immunogenicity of the nonconjugated CP, a more than fivefold increase in the antibody levels was observed when these adjuvants were used with the conjugates. Significant rises in IgG2b and IgG3 were observed with all formulations. The enhancement of the immunogenicity and the IgG subclass shift, as seen with some adjuvants, may prove to be important in immunocompromised patients.

ADP Ribose Transferases↗
PubMed1995

Adjuvant activity of QS-21 for experimental E. coli 018 polysaccharide vaccines.

Three types of experimental vaccines containing O-side-chain polysaccharide from the enterotoxigenic strain Escherichia coli 018 were evaluated. The immunogenicity of free O-polysaccharide (PS), a polysaccharide-diphtheria toxoid conjugate (PS-conj), and detoxified lipopolysaccharide (dLPS) was tested in female ICR mice, either alone or in combination with QS-21, a purified saponin adjuvant derived from the bark of the tree Quillaja saponaria Molina. Both the number of individual mice responding and the titres of O-polysaccharide specific antibodies in pools of sera were increased by the addition of QS-21. The immune response to both O-specific polysaccharide and carrier was primarily IgM and IgG1. The addition of QS-21 not only increased the level of IgG1, but also had a significant adjuvant effect on antigen-specific IgG2a, IgG2b and IgG3.

Adjuvants, Immunologic↗
PubMed1995

Utility of various commercially available human immunodeficiency virus (HIV) antibody diagnostic kits for use in conjunction with efficacy trials of HIV-1 vaccines.

There is a need for human immunodeficiency virus (HIV) screening assays which will distinguish uninfected HIV vaccine recipients from HIV-infected individuals. Commercial screening kits were used to test serum samples from low- and high-risk participants in clinical trials before and after immunization with various recombinant HIV type 1 (HIV-1) envelope glycoprotein 120 (gp120) candidate vaccines. All kits were 100% sensitive in detecting HIV infection. Both Murex Single Use Diagnostic System and United Biomedical, Inc., HIV type 1 or 2 (HIV-1/2) enzyme immunoassay (EIA) kits, which detect antibodies to HIV-1 gp41, were 98 to 100% specific when used to screen baseline or recombinant gp120-vaccinated populations as vaccine-induced antibodies to gp120 were nonreactive in these tests. The Abbott HIVAB HIV-1 EIA (lysate of whole infected cells, reactive with anti-gp120 antibodies) gave high levels of reactivity due to vaccine-induced antibodies and a high baseline rate of false positives (12 of 83) among nonvaccinated high-risk volunteers. Assays containing only gp41 and p24 solid-phase components are compatible with gp120-based vaccines but are unlikely to be useful in a similar role for vaccines containing gp160, gp41, or gp120 plus p24 antigens. Efficacy trials must be designed in concert with available diagnostic screening assays to avoid problems caused by vaccine-induced seroconversion in high-risk populations.

AIDS Vaccines↗
PubMed1995

Time course of Fos-like immunoreactivity associated with cholinergically induced REM sleep.

Now that the pharmacology and neuronal connectivity underlying REM sleep is beginning to be understood, it is important to begin investigations that elucidate the transcriptional response related to the REM sleep process. The present study focuses on determining the temporal development of Fos-like immunoreactivity (Fos-LI) in the dorsolateral pons following cholinergically induced, sustained rapid eye movement (REMc) sleep in cats. Microinjections (0.25 microliter) of vehicle (n = 3) or carbachol (0.2-4.0 micrograms/0.25 microliters) were made into the medial pontine reticular formation. Carbachol produced a state with all the signs of natural REM sleep, and with durations from 0 min to 120 min. Animals were killed either immediately or at various intervals after the end of REMc. Compared to vehicle- and carbachol-treated animals without REMc, the animals with REMc showed a significantly higher number of Fos-LI cells in pontine regions that have been implicated in REM sleep generation. Regions with REMc-associated Fos-LI increases included the lateral dorsal tegmental (LDT) and pedunculopontine tegmental (PPT) nuclei; the locus coeruleus; the dorsal raphe; and the medial pontine reticular formation. More Fos-LI cells were found with longer REMc bouts than with shorter-duration REMc bouts. However, with 2 hr long REMc bouts the number of Fos-LI cells returned to control levels, suggesting that the c-fos transcriptional cascade is turned off once a threshold of REMc has been reached. These findings indicate that pontine neuronal populations implicated in REM sleep express more c-fos in the course of REMc, and that the extent of expression is related to the duration of the state.

Animals↗
PubMed1992

Evaluation of a urease-based confirmatory enzyme-linked immunosorbent assay for diagnosis of Neisseria gonorrhoeae.

A new urease-based enzyme-linked immunosorbent assay utilizing novel monoclonal antibodies was evaluated for the culture confirmation of Neisseria gonorrhoeae, with 270 isolates of N. gonorrhoeae, 56 isolates of diverse Neisseria spp., and 29 Moraxella isolates. The test was highly specific (100.00%) and sensitive (97.83%). No cross-reactions were observed with any of the Neisseria or Moraxella isolates tested. Fifty percent (3 of 6) of the false-negative results were obtained with isolates of serovar IA-4, a serovar rarely encountered in North America.

Antibodies, Monoclonal↗
PubMed1990

The 75-kilodalton protein of Chlamydia trachomatis: a member of the heat shock protein 70 family?

The gene encoding a 75-kilodalton (kDa) protein of Chlamydia trachomatis was cloned, expressed, and sequenced. Genomic libraries from C. trachomatis serovar D DNA were constructed in vectors pUC18 and lambda gt11 and were screened with a panel of monoclonal antibodies against C. trachomatis antigens. The only recombinants identified were those that reacted with antibody UM-13, which has specificity for a genus-specific epitope on the 75-kDa protein. The gene was localized to a 2.9-kilobase DNA fragment and sequenced. The gene consists of a long open reading frame of 1,956 nucleotides, which translates into 652 amino acids totalling 70,558 daltons in mass. Putative promoter elements and a ribosome binding site were identified within 5'-flanking sequences, and a typical rho-independent terminator was identified within 3'-flanking sequences. Screening of the GenBank nucleic acid sequence data bank revealed extensive similarity between the chlamydial 75-kDa gene and the heat shock protein 70 (hsp70) family or proteins. In particular, 71 and 69% amino acid sequence similarities were identified with hsp70 of Escherichia coli and Bacillus megaterium, respectively. Polyclonal antibodies were produced to the recombinant antigen in rabbits and detected epitopes on elementary bodies in enzyme-linked immunosorbent and indirect microimmunofluorescence assays. Antibodies reacted with an antigen of identical molecular mass in L2 and C serovars in an immunoblot assay and neutralized these serovars in cell culture. The 75-kDa protein appears to be a chlamydial homolog of hsp70, is immunoaccessible on native elementary bodies, and is a target for neutralization.

Amino Acid Sequence↗
PubMed1990

Long-term results of catheter ablation of idiopathic right ventricular tachycardia.

Ten consecutive patients with recurrent episodes of symptomatic, idiopathic, sustained monomorphic ventricular tachycardia (VT) originating in the right ventricle underwent an attempt at catheter ablation of the ventricular tachycardia. There were seven women and three men, with a mean age of 39 +/- 14 years (+/- SD). None of the patients had any evidence of structural heart disease. The VT had a left bundle branch block configuration and an inferior axis in each patient, and the mean cycle length was 313 +/- 75 msec. Based on the methods of induction of the VT and the response of the VT to verapamil, the VT mechanism was presumed to be reentry in six patients, triggered activity in three patients, and catecholamine-sensitive automaticity in one patient. Sites for ablation were guided by pace mapping, and an appropriate target site was identified in the right ventricular outflow tract in each patient. From one to three shocks of 100-360 J (mean total, 336 +/- 195 J) were delivered from a defibrillator between the tip of the ablation catheter (cathode) and a patch electrode on the anterior chest (anode). An electrophysiology test 7-9 days after ablation demonstrated that VT was still inducible in only one patient, who was treated with amiodarone. One other patient had a recurrence of VT 3 weeks after ablation and was treated with verapamil. Eight of 10 patients were not treated with antiarrhythmic medications and have had no episodes of symptomatic VT during 15-68 months of follow-up (mean follow-up, 33 +/- 18 months). There were no acute or long-term complications.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1989

The scintigraphic characteristics of ventricular pre-excitation through Mahaim fibers with the use of phase analysis.

The phase image pattern of blood pool scintigrams was blindly assessed in 11 patients exhibiting conduction through Mahaim pathways, including 6 nodoventricular and 5 fasciculoventricular. These patterns were compared with the phase image findings in normal subjects, patients with left and right bundle branch block in the absence of pre-excitation and patients with pre-excitation through atrioventricular (AV) connections. In all patients with a Mahaim pathway, the site of earliest phase angle was septal or paraseptal. Phase progression was asymmetric and the pre-excited ventricle demonstrated the earliest mean ventricular phase angle in 10 of 11 patients. This pattern, and the associated ventricular phase difference, appeared to vary from that in normal subjects and in those with a septal AV connection, in whom phase progression is generally symmetric. Scintigraphic phase analysis provided localizing information and presented patterns consistent with Mahaim pathways. Although not able to differentiate among Mahaim pathway subtypes, these phase patterns differed from those in normal subjects, those with right and left lateral free wall pathways and most patients with a septal AV pathway. However, the phase pattern of patients with a Mahaim pathway may not differ from that of patients with a septal AV connection displaying an asymmetric pattern of phase progression, or those with left and right bundle branch block in the absence of pre-excitation. Objective, yet imperfect phase measurements supported these differences. Such image findings may complement the often complex electrophysiologic evaluation of patients presenting with pre-excitation.

Adult↗
PubMed1989

Detection of Neisseria gonorrhoeae by dot-enzyme immunoassay using monoclonal antibodies.

A highly sensitive and specific dot-enzyme immunoassay for the detection of Neisseria gonorrhoeae was developed using a pool of monoclonal antibodies (MAbs). The MAbs were obtained following immunization of mice with lithium acetate extracted outer membrane (OM) preparations. Western immunoblotting experiments demonstrated that MAbs NG26 and NG38, both IgG2a, reacted with lipopolysaccharides (LPS) and with the major OM protein, P1, respectively, MAb NG28, an IgG3, did not react in Western immunoblotting, MAbs NG28 and NG38 failed to react with OM treated with proteolytic enzymes or with semi-purified preparation of LPS. MAb NG26 reacted with the same LPS preparation. Binding radioimmunoassay with live bacteria showed that all the MAbs adsorbed to cell surface-exposed antigenic determinants. The limit of detection of the dot-enzyme immunoassay was between 1 and 4 x 10(4) cfu per dot. Using a panel of 177 strains of N. gonorrhoeae, MAbs NG28 and NG38 recognized only P1A and P1B strains respectively. MAb NG26 reacted with P1A, P1B and non-typable strains. These MAbs did not react with other Neisseria species or other bacterial species. Using this pool, the dot-enzyme immunoassay had a sensitivity of 93.2% and a specificity of 100%.

Antibodies, Bacterial↗
PubMed1988

Comparison of Visuwell enzyme immunoassay to culture for detection of group A Streptococcus in throat swab specimens.

A microwell enzyme immunoassay (Visuwell) for direct detection of Group A streptococcal antigen from throat swab specimens has been developed. It incorporates urease conjugated antibody as the detector and is easily interpreted by a yellow to purple color change. Throat specimens obtained on rayon-tipped swabs were transported moist in modified Stuarts medium and cultured before being tested in Visuwell (n = 585, prevalence 17.1%, sensitivity 88%, specificity 92.4%, predictive value positive 70.4%, predictive value negative 97.4%, and accuracy 91.6%). In instances of discrepancy between culture and Visuwell, throat swab extracts were tested in a latex agglutination test. In 21 of 37 instances of Visuwell-positive, culture-negative specimens, latex agglutination was positive. Throat specimens obtained using double rayon swabs and transported to the laboratory dry had one swab cultured and the other tested in Visuwell (n = 280, prevalence 20.4%, sensitivity 75.4%, specificity 88.3%, predictive value positive 62.3%, predictive value negative 93.4%, and accuracy 85.7%). When 1+ culture positive specimens were considered negative, a sensitivity of 97.6% was obtained. In 14 of 26 instances of Visuwell-positive, culture-negative specimens, latex agglutination was positive. Cross-reaction with organisms other than Group A Streptococcus found in the oropharynx was negligible in Visuwell. Limit of detection of Group A streptococcal antigen was equivalent for Visuwell and latex agglutination.

Adolescent↗
PubMed1988

Relaxation of DNA torsional tension in defined domains of bacterial chromosomes in vivo.

A procedure is described for selectively relaxing the DNA torsional tension in defined regions of the chromosome of living bacterial cells. Regions of the chromosomal DNA labelled with bromodeoxyuridine are selectively nicked by irradiation of the cells with long-wavelength ultraviolet light and then trimethylpsoralen residues are photobound to the chromosome in vivo. It is demonstrated that the rate of photobinding to the bromouridine-labelled parts of the chromosomes declines relative to the unlabelled parts of the same chromosomes as nicks are introduced into the former regions. The maximal difference in photobinding rates is that expected for the difference between relaxed and negatively supercoiled DNA. Analysis of the number of DNA breaks required for minimizing the photobinding rates permits a calculation of the number of domains of supercoiling per Bacillus subtilis chromosome.

Bacillus subtilis↗