We have attempted to simplify the procedure for coupling various ligands to distal ends of liposome-grafted polyethylene glycol (PEG) chains and to make it applicable for single-step binding of a large variety of a primary amino group-containing substances, including proteins and small molecules. With this in mind, we have introduced a new amphiphilic PEG derivative, p-nitrophenylcarbonyl-PEG-1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (pNP-PEG-DOPE), synthesized by reaction of DOPE with excess of bis(p-nitrophenylcarbonyl)-PEG in a chloroform/triethylamine mixture. pNP-PEG-DOPE readily incorporates into liposomes via its PE residue, and easily binds primary amino group-containing ligands via its water-exposed pNP groups, forming stable and non-toxic urethane (carbamate) bonds. The reaction between the pNP group and the ligand amino group proceeds easily and quantitatively at pH around 8.0, and remaining free pNP groups are promptly eliminated by spontaneous hydrolysis. Therefore, pNP-PEG-DOPE could serve as a very convenient tool for protein attachment to the distal ends of liposome-grafted PEG chains. To investigate the applicability of the suggested protocol for the preparation of long-circulating targeted liposomes, we have coupled several proteins, such as concanavalin A (ConA), wheat germ agglutinin (WGA), avidin, monoclonal antimyosin antibody 2G4 (mon2G4), and monoclonal antinucleosome antibody 2C5 (mon2C5) to PEG-liposomes via terminal pNP groups and studied whether the specific activity of these immobilized proteins is preserved. The method permits the binding of several dozens protein molecules per single 200 nm liposome. All bound proteins completely preserve their specific activity. Lectin-liposomes are agglutinated by the appropriate polyvalent substrates (mannan for ConA-liposomes and glycophorin for WGA-liposomes); avidin-liposomes specifically bind with biotin-agarose; antibody-liposomes demonstrate high specific binding to the substrate monolayer both in the direct binding assay and in ELISA. A comparison of the suggested method with the method of direct membrane incorporation was made. The effect of the concentration of liposome-grafted PEG on the preservation of specific protein activity in different coupling protocols was also investigated. It was also shown that pNP-PEG-DOPE-liposomes with and without attached ligands demonstrate increased stability in mouse serum.

NGF has been suggested to play a role in urinary bladder dysfunction by mediating inflammation, as well as morphological and functional changes, in sensory and sympathetic neurons innervating the urinary bladder. To further explore the role of NGF in bladder sensory function, we generated a transgenic mouse model of chronic NGF overexpression in the bladder using the urothelium-specific uroplakin II (UPII) promoter. NGF mRNA and protein were expressed at higher levels in the bladders of NGF-overexpressing (NGF-OE) transgenic mice compared with wild-type littermate controls from postnatal day 7 through 12-16 wk of age. Overexpression of NGF led to urinary bladder enlargement characterized by marked nerve fiber hyperplasia in the submucosa and detrusor smooth muscle and elevated numbers of tissue mast cells. There was a marked increase in the density of CGRP- and substance P-positive C-fiber sensory afferents, neurofilament 200-positive myelinated sensory afferents, and tyrosine hydroxylase-positive sympathetic nerve fibers in the suburothelial nerve plexus. CGRP-positive ganglia were also present in the urinary bladders of transgenic mice. Transgenic mice had reduced urinary bladder capacity and an increase in the number and amplitude of nonvoiding bladder contractions under baseline conditions in conscious open-voiding cystometry. These changes in urinary bladder function were further associated with an increased referred somatic pelvic hypersensitivity. Thus, chronic urothelial NGF overexpression in transgenic mice leads to neuronal proliferation, focal increases in urinary bladder mast cells, increased urinary bladder reflex activity, and pelvic hypersensitivity. NGF-overexpressing mice may, therefore, provide a useful transgenic model for exploring the role of NGF in urinary bladder dysfunction.

Fibromyalgia is the diagnosis given to individuals with chronic widespread musculoskeletal pain for which no alternative cause, such as tissue inflammation or damage, can be identified. Fibromyalgia is now believed to be, at least in part, a disorder of central pain processing that produces heightened responses to painful stimuli (hyperalgesia) and painful responses to nonpainful stimuli (allodynia). Aberrations in central pain processing may also be partly responsible for symptoms experienced in several chronic pain disorders that coaggregate with fibromyalgia, which is itself a product of genetic and environmental factors. Thus, aberrational central pain processing is implicated in irritable bowel syndrome, temporomandibular disorder, chronic low back pain, and certain other chronic pain disorders. Fibromyalgia and related disorders appear to reflect deficiencies in serotonergic and noradrenergic, but not opioidergic, transmission in the central nervous system. The heightened state of pain transmission may also be owing to increases in pronociceptive neurotransmitters such as glutamate and substance P. In some cases, psychological and behavioral factors are also in play. Although the overlapping symptomatology between fibromyalgia and related disorders may present diagnostic challenges, proper examination and observation can help clinicians make an accurate diagnosis. In recent years, the vastly improved understanding of the mechanism underlying fibromyalgia and the related spectrum of diseases has fostered rapid advances in the therapy of these chronic pain disorders by both pharmacologic and nonpharmacologic interventions.

BACKGROUND

Berries are a particularly rich source of polyphenols. They also contain other bioactive substances, such as vitamin C. Previous studies indicated that the consumption of polyphenol-rich foods (eg, cocoa, tea, and red wine) may induce beneficial changes in pathways related to cardiovascular health. Whether the consumption of berries has similar effects is unknown.

OBJECTIVE

We aimed to investigate the effects of berry consumption on hemostatic function, serum lipids, and blood pressure (BP).

METHODS

Middle-aged unmedicated subjects (n = 72) with cardiovascular risk factors consumed moderate amounts of berry or control products for 8 wk in a single-blind, randomized, placebo-controlled intervention trial.

RESULTS

Berry consumption inhibited platelet function as measured with a platelet function analyzer (using collagen and ADP as platelet activator) [changes: 11% and -1.4% in the berry and control groups, respectively; P = 0.018, analysis of covariance (ANCOVA)]. Plasma biomarkers of platelet activation, coagulation, and fibrinolysis did not change during the intervention. Serum HDL-cholesterol concentrations increased significantly more (P = 0.006, ANCOVA) in the berry than in the control group (5.2% and 0.6%, respectively), but total cholesterol and triacylglycerol remained unchanged. Systolic BP decreased significantly (P = 0.050, ANCOVA); the decrease mostly occurred in subjects with high baseline BP (7.3 mm Hg in highest tertile; P = 0.024, ANCOVA). Polyphenol and vitamin C concentrations in plasma increased, whereas other nutritional biomarkers (ie, folate, tocopherols, sodium, and potassium) were unaffected.

CONCLUSIONS

The consumption of moderate amounts of berries resulted in favorable changes in platelet function, HDL cholesterol, and BP. The results indicate that regular consumption of berries may play a role in the prevention of cardiovascular disease.

In light of the possible role peripheral nerves may play in bone metabolism, the morphology of calcitonin gene-related peptide (CGRP)-, vasoactive intestinal peptide (VIP)-, substance P (SP)-, neuropeptide Y (NPY)-, and dopamine-beta-hydroxylase (D beta H)-immunoreactive nerve fibers was examined in whole-mount preparations of periosteum of membranous bones (calvaria, mandible) and long bones (tibia) from the rat. Periosteum from animals treated to remove selectively either the sympathetic or fine-caliber primary afferent nerves was also examined to determine the origin of the nerve fibers. We found a consistent and often dense innervation of the periosteum. The innervation patterns of the calvaria and mandible were similar, with networks of nerves spread across the surface of the bone. Nerves in the tibial periosteum were oriented in the longitudinal axis and were more numerous at the epiphyses than in the mid-shaft region. CGRP-immunoreactive fibers were widely and densely distributed. The presence of populations of CGRP-immunoreactive fibers of differing calibers and perivascular arrangements suggests that such nerves in bone tissues may serve different functions. SP-immunoreactivity was present in a fine network of varicose fibers in the superficial layers of the periosteum. CGRP- and SP-immunoreactive nerve fibers were dramatically reduced in periosteum of capsaicin-treated animals as compared to controls, indicating the sensory origin of these nerves. VIP-immunoreactive nerve fibers were distributed in the periosteum of mandible and calvaria as small networks and individual fine varicose fibers. In tibial periosteum, larger networks of these fibers were visible. VIP-immunoreactive nerve fibers in the periosteum were associated with both vascular and nonvascular elements within the layers of cells closest to the bone, suggesting that VIP may serve more than one function in periosteal tissues. NPY-immunoreactive fibers were largely confined to vascular elements; occasional fibers were observed among the bone-lining cells. D beta H-immunoreactivity was associated only with blood vessels. VIP-, NPY-, and D beta H-immunoreactivities were dramatically reduced in the periosteum of guanethidine-treated animals, indicating the sympathetic origin of these nerves.

BACKGROUND

Selective interventions targeting personality risk are showing promise in the prevention of problematic drinking behavior, but their effect on illicit drug use has yet to be evaluated.

OBJECTIVE

To investigate the efficacy of targeted coping skills interventions on illicit drug use in adolescents with personality risk factors for substance misuse.

METHODS

Randomized controlled trial.

METHODS

Secondary schools in London, United Kingdom.

METHODS

A total of 5302 students were screened to identify 2028 students aged 13 to 16 years with elevated scores on self-report measures of hopelessness, anxiety sensitivity, impulsivity, and sensation seeking. Seven hundred thirty-two students provided parental consent to participate in this trial.

METHODS

Participants were randomly assigned to a control no-intervention condition or a 2-session group coping skills intervention targeting 1 of 4 personality profiles.

METHODS

The trial was designed and powered to primarily evaluate the effect of the intervention on the onset, prevalence, and frequency of illicit drug use over a 2-year period.

RESULTS

Intent-to-treat repeated-measures analyses on continuous measures of drug use revealed time x intervention effects on the number of drugs used (P < .01) and drug use frequency (P < .05), whereby the control group showed significant growth in the number of drugs used as well as more frequent drug use over the 2-year period relative to the intervention group. Survival analysis using logistic regression revealed that the intervention was associated with reduced odds of taking up the use of marijuana (beta = -0.3; robust SE = 0.2; P = .09; odds ratio = 0.7; 95% confidence interval, 0.5-1.0), cocaine (beta = -1.4; robust SE = 0.4; P < .001; odds ratio = 0.2; 95% confidence interval, 0.1-0.5), and other drugs (beta = -0.7; robust SE = 0.3; P = .03; odds ratio = 0.5; 95% confidence interval, 0.3-0.9) over the 24-month period.

CONCLUSIONS

This study extends the evidence that brief, personality-targeted interventions can prevent the onset and escalation of substance misuse in high-risk adolescents.

BACKGROUND

clinicaltrials.gov Identifier: NCT00344474.

BACKGROUND

Personality factors are implicated in the vulnerability to adolescent alcohol misuse. This study examined whether providing personality-targeted interventions in early adolescence can delay drinking and binge drinking in high-risk youth.

METHODS

A randomised control trial was carried out with 368 adolescents recruited from years 9 and 10 (median age 14) with personality risk factors for substance misuse. Participants received either a personality-targeted intervention or no intervention. Outcome data were collected on alcohol use through self-reports at 6 and 12-month post intervention and analyses were conducted on the full intent to treat sample.

RESULTS

Multi-group analysis of a latent growth curve model showed a group difference in the growth of alcohol use between baseline and 6-months follow-up, with the control group showing a greater increase in drinking than the intervention group for this period. Interventions were particularly effective in preventing the growth of binge drinking in those students with a sensation seeking (SS) personality. SS drinkers in the intervention group were 45% and 50% less likely to binge drink at 6 (OR = .45) and 12 months (OR = .50) respectively, than SS drinkers in the control group, p = .001, phi = .49, Number Needed to Treat = 2.0.

CONCLUSIONS

Considering the robust, inverse relationship between age of onset of alcohol use and later alcohol dependence, this intervention strategy may prove effective in preventing the onset of adult alcohol use disorders, by helping high-risk youth delay the growth of their drinking to a later developmental stage.

OBJECTIVE

This study aimed to determine the incidence of psychiatric disorders among health care workers in Toronto in the one- to two-year period after the 2003 outbreak of severe acute respiratory syndrome (SARS) and to test predicted risk factors.

METHODS

New-onset episodes of psychiatric disorders were assessed among 139 health care workers by using the Structured Clinical Interview for DSM-IV and the Clinician-Administered PTSD Scale. Past history of psychiatric illness, years of health care experience, and the perception of adequate training and support were tested as predictors of the incidence of new-onset episodes of psychiatric disorders after the SARS outbreak.

RESULTS

The lifetime prevalence of any depressive, anxiety, or substance use diagnosis was 30%. Only one health care worker who identified the SARS experience as a traumatic event was diagnosed as having PTSD. New episodes of psychiatric disorders occurred among seven health care workers (5%). New episodes of psychiatric disorders were directly associated with a history of having a psychiatric disorder before the SARS outbreak (p=.02) and inversely associated with years of health care experience (p=.03) and the perceived adequacy of training and support (p=.03).

CONCLUSIONS

Incidence of new episodes of psychiatric disorders after the SARS outbreak were similar to or lower than community incidence rates, which may indicate the resilience of health care workers who continued to work in hospitals one to two years after the SARS outbreak. In preparation for future events, such as pandemic influenza, training and support may bolster the resilience of health care workers who are at higher risk by virtue of their psychiatric history and fewer years of health care experience.

Extensive evidence indicates that disruption of cholinergic function is characteristic of aging and Alzheimer's disease (AD), and experimental manipulation of the cholinergic system in laboratory animals suggests age-related cholinergic dysfunction may play an important role in cognitive deterioration associated with aging and AD. Recent research, however, suggests that cholinergic dysfunction does not provide a complete account of age-related cognitive deficits and that age-related changes in cholinergic function typically occur within the context of changes in several other neuromodulatory systems. Evidence reviewed in this paper suggests that interactions between the cholinergic system and several of these neurotransmitters and neuromodulators--including norepinephrine, dopamine, serotonin, GABA, opioid peptides, galanin, substance P, and angiotensin II--may be important in learning and memory. Thus, it is important to consider not only the independent contributions of age-related changes in neuromodulatory systems to cognitive decline, but also the contribution of interactions between these systems to the learning and memory deficits associated with aging and AD.

The sympathetic and sensory innervation of guinea-pig trachea and lung were studied by means of retrograde neuronal tracing using fluorescent dyes, and double-labelling immunofluorescence. Sympathetic neurons supplying the lung were located in stellate ganglia and in thoracic sympathetic chain ganglia T2-T4; those supplying the trachea resided in the superior cervical and stellate ganglia. Retrogradely labelled sympathetic neurons were usually immunoreactive to tyrosine hydroxylase; the majority also contained neuropeptide Y immunoreactivity. However, a small number were non-catecholaminergic (i.e. tyrosine hydroxylase negative), but neuropeptide Y immunoreactive. Within the airways, tyrosine hydroxylase/neuropeptide Y-immunoreactive axons were found in the smooth muscle layer, around blood vessels including the pulmonary artery and vein, and to a lesser extent in the lamina propria. Periarterial axons contained in addition dynorphin immunoreactivity. Sensory neurons supplying the lung were located in jugular and nodose vagal ganglia as well as in upper thoracic dorsal root ganglia; those supplying the trachea were most frequently found bilaterally in the nodose ganglia and less frequently in the jugular ganglia. A spinal origin of tracheal sensory fibres could not be consistently demonstrated. With regard to their immunoreactivity to peptides, three types of sensory neurons projecting to the airways could be distinguished: (i) substance P/dynorphin immunoreactive; (ii) substance P immunoreactive but dynorphin negative; and (iii) negative to all peptides tested. Substance P-immunoreactive neurons innervating the airways invariably contained immunoreactivity to neurokinin A and calcitonin gene-related peptide. Retrogradely labelled neurons located in the nodose ganglia belonged almost exclusively (greater than or equal to 99%) to the peptide-negative group, whereas the three neuron types each represented about one-third of retrogradely labelled neurons in jugular and dorsal root ganglia. Within the airways, axons immunoreactive to substance P/neurokinin A and substance P/calcitonin gene-related peptide were distributed within the respiratory epithelium of trachea and large bronchi, in the lamina propria and smooth muscle from the trachea down to the smallest bronchioli (highest density at the bronchial level), in the alveolar walls, around systemic and pulmonary blood vessels, and within airway ganglia. Those axons also containing dynorphin immunoreactivity were restricted to the lamina propria and smooth muscle. The origin of nerve fibres immunoreactive for vasoactive intestinal polypeptide, of which a part were also neuropeptide Y immunoreactive, could not be determined by retrograde tracing experiments. Vasoactive intestinal polypeptide-immunoreactive fibres terminating within airway ganglia may be of preganglionic parasympathetic origin, whereas others (e.g. those found in smooth muscle) may arise from intrinsic ganglia.(ABSTRACT TRUNCATED AT 400 WORDS)

For both type 1 and type 2 diabetes mellitus, the rates have been increasing in the United States and elsewhere; rates vary widely by country, and genetic factors account for less than half of new cases. These observations suggest environmental factors cause both type 1 and type 2 diabetes. Occupational exposures have been associated with increased risk of diabetes. In addition, recent data suggest that toxic substances in the environment, other than infectious agents or exposures that stimulate an immune response, are associated with the occurrence of these diseases. We reviewed the epidemiologic data that addressed whether environmental contaminants might cause type 1 or type 2 diabetes. For type 1 diabetes, higher intake of nitrates, nitrites, and N-nitroso compounds, as well as higher serum levels of polychlorinated biphenyls have been associated with increased risk. Overall, however, the data were limited or inconsistent. With respect to type 2 diabetes, data on arsenic and 2,3,7,8-tetrachlorodibenzo-p-dioxin relative to risk were suggestive of a direct association but were inconclusive. The occupational data suggested that more data on exposure to N-nitroso compounds, arsenic, dioxins, talc, and straight oil machining fluids in relation to diabetes would be useful. Although environmental factors other than contaminants may account for the majority of type 1 and type 2 diabetes, the etiologic role of several contaminants and occupational exposures deserves further study.

Elevated extracellular K(+) ([K(+)](o)), in the absence of "classical" immunological stimulatory signals, was found to itself be a sufficient stimulus to activate T cell beta1 integrin moieties, and to induce integrin-mediated adhesion and migration. Gating of T cell voltage-gated K(+) channels (Kv1.3) appears to be the crucial "decision-making" step, through which various physiological factors, including elevated [K(+)](o) levels, affect the T cell beta1 integrin function: opening of the channel leads to function, whereas its blockage prevents it. In support of this notion, we found that the proadhesive effects of the chemokine macrophage-inflammatory protein 1beta, the neuropeptide calcitonin gene-related peptide (CGRP), as well as elevated [K(+)](o) levels, are blocked by specific Kv1.3 channel blockers, and that the unique physiological ability of substance P to inhibit T cell adhesion correlates with Kv1.3 inhibition. Interestingly, the Kv1.3 channels and the beta1 integrins coimmunoprecipitate, suggesting that their physical association underlies their functional cooperation on the T cell surface. This study shows that T cells can be activated and driven to integrin function by a pathway that does not involve any of its specific receptors (i.e., by elevated [K(+)](o)). In addition, our results suggest that undesired T cell integrin function in a series of pathological conditions can be arrested by molecules that block the Kv1.3 channels.

BACKGROUND

Although the basal ganglia are thought to play a key role in action selection in mammals, it is unknown whether this mammalian circuitry is present in lower vertebrates as a conserved selection mechanism. We aim here, using lamprey, to elucidate the basal ganglia circuitry in the phylogenetically oldest group of vertebrates (cyclostomes) and determine how this selection architecture evolved to accommodate the increased behavioral repertoires of advanced vertebrates.

RESULTS

We show, using immunohistochemistry, tract tracing, and whole-cell recordings, that all parts of the mammalian basal ganglia (striatum, globus pallidus interna [GPi] and externa [GPe], and subthalamic nucleus [STN]) are present in the lamprey forebrain. In addition, the circuit features, molecular markers, and physiological activity patterns are conserved. Thus, GABAergic striatal neurons expressing substance P project directly to the pallidal output layer, whereas enkephalin-expressing striatal neurons project indirectly via nuclei homologous to the GPe and STN. Moreover, pallidal output neurons tonically inhibit tectum, mesencephalic, and diencephalic motor regions.

CONCLUSIONS

These results show that the detailed basal ganglia circuitry is present in the phylogenetically oldest vertebrates and has been conserved, most likely as a mechanism for action selection used by all vertebrates, for over 560 million years. Our data also suggest that the mammalian basal ganglia evolved through a process of exaptation, where the ancestral core unit has been co-opted for multiple functions, allowing them to process cognitive, emotional, and motor information in parallel and control a broader range of behaviors.

Increasing evidence implicates the substance P (SP)/neurokinin-1 receptor system in anxiety and depression. However, it is not known whether emotional stimulation alters endogenous extracellular SP levels in brain areas important for processing of anxiety and mood, a prerequisite for a contribution of this neuropeptide system in modulating these behaviors. Therefore, we examined in rats whether the release of SP is sensitive to emotional stressors in distinct subregions of the amygdala, a key area in processing of emotions. By using in vivo micropush-pull superfusion and microdialysis techniques, we found a pronounced and long-lasting increase (150%) in SP release in the medial nucleus of the amygdala (MeA), but not in the central nucleus of the amygdala, in response to immobilization stress. SP release in the MeA was transiently enhanced (40%) in response to elevated platform exposure, which is regarded as a mild emotional stressor. Immobilization enhanced the anxiety-related behavior evaluated in the subsequently performed elevated plus-maze test. Bilateral microinjections of the neurokinin-1 receptor antagonist [2-cyclopropoxy-5-(5-(trifluoromethyl)tetrazol-1-yl)benzyl]-(2-phenylpiperidin-3-yl)amine into the MeA blocked the stress-induced anxiogenic-like effect, supporting a functional significance of enhanced SP release. In unstressed rats, the neurokinin-1 receptor antagonist displayed no significant anxiolytic effect but reversed the anxiogenic effect of SP microinjected into the MeA. Our findings identify the MeA as a critical brain area for the involvement of SP transmission in anxiety responses and as a putative site of action for the recently discovered therapeutic effects of SP antagonists in the treatment of stress-related disorders.

The present study was designed to assess the influence of deviant peer affiliations on crime and substance use in adolescence/young adulthood. Data were used from a 21-year longitudinal study of health, development, and adjustment of a birth cohort of 1,265 New Zealand children. Annual assessments of deviant peer affiliations were obtained for the period from age 14-21 years, together with measures of psychosocial outcomes including, violent crime, property crime, alcohol abuse, cannabis abuse, and nicotine dependence. Affiliating with deviant peers was found to be significantly associated with each of these outcomes (p < .0001). Statistical control for confounding by both fixed and time dynamic factors reduced the strength of association between deviant peer affiliations and outcome measures. Nevertheless, deviant peer affiliations remained significantly associated (p < .0001) with all outcomes. For violent/property crime, cannabis and alcohol abuse there was significant evidence of age-related variation in the strength of association with deviant peer affiliations, with deviant peer affiliations having greater influence on younger participants (14-15 years) than older participants (20-21 years). These results suggest that deviant peer affiliations are associated with increased rates of a range of adjustment problems in adolescence/young adulthood with deviant peer affiliations being most influential at younger ages.

BACKGROUND

Social and communication impairments are core deficits and prognostic indicators of autism. We evaluated the impact of supplementing a comprehensive intervention with a curriculum targeting socially synchronous behavior on social outcomes of toddlers with autism spectrum disorders (ASD).

METHODS

Fifty toddlers with ASD, ages 21 to 33 months, were randomized to one of two six-month interventions: Interpersonal Synchrony or Non-Interpersonal Synchrony. The interventions provided identical intensity (10 hours per week in classroom), student-to-teacher ratio, schedule, home-based parent training (1.5 hours per month), parent education (38 hours), and instructional strategies, except the Interpersonal Synchrony condition provided a supplementary curriculum targeting socially engaged imitation, joint attention, and affect sharing; measures of these were primary outcomes. Assessments were conducted pre-intervention, immediately post-intervention, and, to assess maintenance, at six-month follow-up. Random effects models were used to examine differences between groups over time. Secondary analyses examined gains in expressive language and nonverbal cognition, and time effects during the intervention and follow-up periods.

RESULTS

A significant treatment effect was found for socially engaged imitation (p = .02), with more than doubling (17% to 42%) of imitated acts paired with eye contact in the Interpersonal Synchrony group after the intervention. This skill was generalized to unfamiliar contexts and maintained through follow-up. Similar gains were observed for initiation of joint attention and shared positive affect, but between-group differences did not reach statistical significance. A significant time effect was found for all outcomes (p < .001); greatest change occurred during the intervention period, particularly in the Interpersonal Synchrony group.

CONCLUSIONS

This is the first ASD randomized trial involving toddlers to identify an active ingredient for enhancing socially engaged imitation. Adding social engagement targets to intervention improves short-term outcome at no additional cost to the intervention. The social, language, and cognitive gains in our participants provide evidence for plasticity of these developmental systems in toddlers with ASD. http://www.clinicaltrials.gov/ct2/show/NCT00106210?term = landa&rank = 3.

The role of peptides as signalling molecules in the nervous system has been studied for more than 30 years. Neuropeptides and their G-protein-coupled receptors are widely distributed throughout the body and they commonly occur with, and are complementary to, classic neurotransmitters. The functions of neuropeptides range from neurotransmitter to growth factor. They are present in glial cells, are hormones in the endocrine system, and are messengers in the immune system. Much evidence indicates that neuropeptides are of particular importance when the nervous system is challenged (eg, by stress, injury, or drug abuse). These features and the large number of neuropeptides and neuropeptide receptors provide many opportunities for the discovery of new drug targets for the treatment of nervous-system disorders. In fact, receptor-subtype-selective antagonists and agonists have been developed, and recently a substance P receptor (neurokinin 1) antagonist has been shown to have clinical efficacy in the treatment of major depression and chemotherapy-induced emesis. Several other neuropeptide receptor ligands are in clinical trials for various indications.

Previous studies have demonstrated the existence of a circulating myocardial depressant substance during human septic shock. We have recently identified this substance as a synergistic combination of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta). This study utilized an in vitro cardiac myocyte assay to evaluate the potential mechanistic role of nitric oxide (NO) and cGMP in depression of myocyte contractility induced by TNF-alpha, IL-1beta, TNF-alpha + IL-1beta (at low concentrations), and human septic shock serum (HSS). TNF-alpha, IL-1beta, TNF-alpha + IL-1beta, and each of 5 sera from patients with acute septic shock caused depression of both maximum extent and peak velocity of cardiac myocyte shortening and an increase in intracellular cGMP concentration during 30 min of exposure (minimum P < 0.01). NO synthetase (NOS) and guanylate cyclase inhibitors such as N-methyl-L-arginine (L-NMA) and methylene blue prevented these effects; an excess of L-arginine with L-NMA restored them (minimum P < 0.01). In contrast, D-arginine failed to reestablish cytokine-induced myocyte depression and cGMP accumulation prevented by L-NMA. Exposure of myocytes to TNF-alpha, IL-1beta, or TNF-alpha + IL-1beta produced a concentration-dependent increase in intracellular cGMP that paralleled the depression of cardiac myocyte contractility (minimum P < 0.001). In addition, TNF-alpha, IL-1beta, TNF-alpha + IL-1beta, or HSS application to cardiac myocytes resulted in increased NO gas generation, which was inhibited by L-NMA (minimum P < 0.01). Furthermore, unstimulated cardiac myocytes were shown to harbor constitutive but not inducible NOS activity. These data suggest that the sequential generation of NO by a constitutive NOS and cGMP by guanylate cyclase represents an important mechanism of cardiac myocyte depression by TNF-alpha, IL-1beta, TNF-alpha + IL-1beta, and the myocardial depressant substance(s) of septic shock.

Several sympathetic ganglia of the guinea pig, cat and rat were studied with indirect immunofluorescence technique using antibodies to substance P (SP) and dopamine-beta-hydroxylase (DBH), the latter of which represents a marker for noradrenaline containing neurons. In all ganglia studied SP-like immunoreactivity could be observed in certain nerve fibers (SP-positive fibers). Their number, localization fluorescence intensity and 'fluorescence morphology' varied considerably between the different ganglia as well as between species. In the inferior mesenteric ganglion and the coeliac-superior mesenteric ganglion complex of all species, and in particular of the guinea pig, a dense plexus of varicose SP-positive fibers was observed around the mostly DBH-positive, principal ganglion cells. In contrast no such fibers were seen in relation to the DBH-positive SIF cells of the ganglia. No SP-positive ganglion cells were observed in any of the ganglia studied. Previous biochemical and immunohistochemical studies have indicated that peripheral SP-positive fibers may represent the branches of primary sensory neurons. The rich supply of varicose, SP-positive nerve terminals, often surrounding the noradrenergic ganglion cells in a basket-like manner, may indicate that these fibers may play a functional role within the ganglia. Instead of merely passing through the ganglion they may be involved in special types of reflex arches.

A detailed account of the distribution of immunoreactive substance P-containing structures in the rat central nervous system is presented, from results obtained by applying an indirect immunofluorescent technique. High densities of substance P-containing nerve terminals were present in sensory nuclei and other non-sensory structures such as thalamus, hypothalamus and extrapyramidal system. Substance P-reactive neuron cell bodies were present in spinal root ganglia, nucleus habenulae medialis, nucleus interpeduncularis, caudoputamen and globus pallidus. Most of the neocortex and the cerebellar cortices had no substance P-positive elements. The results support the hypothesis that substance P may be a widespread neurotransmitter in the central nervous system.

Substance P (6.25-25 p-mole) produced dose-dependent flare and wheal responses when injected intradermally into the volar surface of the human forearm. The maximum flare response was obtained within the first 3 min of injection and declined thereafter. The wheal response reached a maximum after 12 min following the injection. Only those peptides having one or more basic residues in the N-terminal region were effective in producing a flare reaction. Eledoisin-related peptide and SPsubstance P respectively, whilst [D-pro2, D-phe7, D-trpPpeptide, SPproducing activity was not dependent on the presence of basic residues and the rank order of relative potencies was: physalaemin (2.0): [D-pro2, D-phe7, D-trpPPPPpeptide (0.08): eledoisin (0.06). The N-terminal tetrapeptide failed to produce a wheal response in the dose range tested. Substance P was approximately equi-active with poly-L-arginine in the production of wheal and flare and both of these agents were about 10 times more potent than histamine. Adenosine triphosphate (25-400 n-mole) produced dose-dependent wheal and flare responses and was 10,000 times less potent than substance P. Pre-treatment of the subjects with the H1 histamine antagonist, chlorpheniramine, (20 mg I.V.) reduced the wheal and flare responses to substance P. Local anaesthetic injection into the skin reduced the spread of the flare response but did not affect the development of the wheal response. Pre-treatment of the skin with capsaicin reduced the flare but not the wheal response to intradermal injection of histamine. The results are discussed in relation to the mechanism of the 'axon reflex' vasodilatation in skin. This is thought to involve mast cells in addition to substance P-containing primary afferent neurones.

Anderson, Douglas R. (National Cancer Institute, Bethesda, Md.), Hope E. Hopps, Michael F. Barile, and Barbara C. Bernheim. Comparison of the ultrastructure of several rickettsiae, ornithosis virus, and Mycoplasma in tissue culture. J. Bacteriol. 90:1387-1404. 1965.-In an effort to make a valid comparison of the ultrastructure of several intracellular parasites, selected agents were propagated under identical conditions in a single type of tissue culture cell; such infected preparations were processed for examination by electron microscopy by use of a standardized procedure for fixation and embedding. The organisms studied were: the Breinl and E strains of epidemic typhus, Rickettsia prowazeki; the Bitterroot strain of R. rickettsii; the Karp strain of R. tsutsugamushi (R. orientalis); R. sennetsu; the P-4 strain of ornithosis virus; and the HEp-2 strain of Mycoplasma hominis type I. Each of the rickettsial species examined had a cell wall and a plasma membrane, and contained ribosomes and deoxyribonucleic acid (DNA) in a ground substance. However, certain differences were noted. Both strains of R. prowazeki contained numerous intracytoplasmic electron-lucent spherical structures (4 to 10 mmu), not previously described. R. sennetsu, unlike the other rickettsiae, was not free in the host cytoplasm but was always enclosed in a vacuole. R. rickettsii was observed intranuclearly and in digestive organelles of the host cell as well as in the cytoplasm. Cells infected with ornithosis virus contained several forms representing the stages in its life cycle. The "initial bodies," made up of ribosomes and DNA strands, were morphologically similar to the rickettsiae. In cultures infected with M. hominis, most of the cells became large and multinucleate. Although the Mycoplasma organisms were readily cultivated from these cultures, only a few could be found in the electron microscope preparations. These organisms were extracellular and lacked a cell wall, being bound only by a unit membrane. Again, the internal components were ribosomes and DNA strands. Under the uniform preparative conditions employed here, the three groups of organisms were morphologically distinguishable from one another.

The presence of calcitonin-gene related peptide (CGRP)-like immunoreactivity (-LI) in sensory neurons was established by immunohistochemistry and radioimmunoassay (RIA) in combination with high performance liquid chromatography (HPLC). CGRP-immunoreactive (-IR) nerve fibres were present in many peripheral organs including heart, ureter, uterus and gall bladder of guinea-pig and man. The distribution of CGRP-IR nerves in the dorsal horn of the spinal cord, of positive cell bodies in thoracic spinal and nodose ganglia and nerves in peripheral organs was closely related to that of substance P-LI. Double staining experiments revealed that in most cases peripheral CGRP-IR nerve terminals also contained SP-LI. However, different localization of SP- and CGRP-IR neurons was observed in the nucleus of the solitary tract as well as in the ventral horn of the spinal cord. In the heart, CGRP-IR nerves were associated with myocardial cells (mainly atria), coronary vessels, local parasympathetic ganglia as well as with the epi- and endocardia. Three to 4-fold higher levels of native CGRP-LI were observed in the atria than in the ventricles of the heart. HPLC analysis revealed that the major peak of CGRP-LI in the heart of rat and man had the same retention times as the synthetic equivalents. Systemic capsaicin pretreatment and adult guinea-pigs caused a loss of CGRP-IR terminals in the dorsal horn of the spinal cord as well as in peripheral organs including the heart. After capsaicin treatment, the content of CGRP-IR was reduced by 70% in the heart and by 60% in the dorsal part of the spinal cord. In superfusion experiments with slices from the rat spinal cord, a release of CGRP-LI was induced by 60 mM K+ and 3 microM capsaicin in a calcium-dependent manner.

BACKGROUND

A dry, persistent cough is a well-described class effect of the angiotensin-converting enzyme (ACE) inhibitor medications. The mechanism of ACE inhibitor-induced cough remains unresolved, but likely involves the protussive mediators bradykinin and substance P, agents that are degraded by ACE and therefore accumulate in the upper respiratory tract or lung when the enzyme is inhibited, and prostaglandins, the production of which may be stimulated by bradykinin.

METHODS

Data for this review were obtained from a National Library of Medicine (PubMed) search, which was performed in May 2004, of the literature published in the English language from 1985 to 2004, using the search terms "angiotensin-converting enzyme," "angiotensin converting enzyme inhibitors," and "cough."

RESULTS

The incidence of ACE inhibitor-induced cough has been reported to be in the range of 5 to 35% among patients treated with these agents. However, a much lower incidence has been described in studies of patients presenting for the evaluation of chronic cough. The onset of ACE inhibitor-induced cough ranges from within hours of the first dose to months after the initiation of therapy. Resolution typically occurs within 1 to 4 weeks after the cessation of therapy, but cough may linger for up to 3 months. The only uniformly effective treatment for ACE inhibitor-induced cough is the cessation of treatment with the offending agent. The incidence of cough associated with therapy with angiotensin-receptor blockers appears to be similar to that of the control drug. In a minority of patients, cough will not recur after the reintroduction of ACE inhibitor therapy.

CONCLUSIONS

In a patient with chronic cough, ACE inhibitors should be considered as wholly or partially causative, regardless of the temporal relation between the initiation of ACE inhibitor therapy and the onset of cough. Although the cessation of therapy is the only uniformly effective treatment for ACE inhibitor-induced cough, some pharmacologic agents have been shown to attenuate the cough.