Maintained intentional weight loss reduces cardiovascular outcomes: results from the Sibutramine Cardiovascular OUTcomes (SCOUT) trial.

AIM: The Sibutramine Cardiovascular OUTcomes trial showed that sibutramine produced greater mean weight loss than placebo but increased cardiovascular morbidity but not mortality. The relationship between 12-month weight loss and subsequent cardiovascular outcomes is explored. METHODS: Overweight/obese subjects (N = 10 744), ≥55 years with cardiovascular disease and/or type 2 diabetes mellitus, received sibutramine plus weight management during a 6-week Lead-in Period before randomization to continue sibutramine (N = 4906) or to receive placebo (N = 4898). The primary endpoint was the time from randomization to first occurrence of a primary outcome event (non-fatal myocardial infarction, non-fatal stroke, resuscitated cardiac arrest or cardiovascular death). RESULTS: For the total population, mean weight change during Lead-in Period (sibutramine) was -2.54 kg. Post-randomization, mean total weight change to Month 12 was -4.18 kg (sibutramine) or -1.87 kg (placebo). Degree of weight loss during Lead-in Period or through Month 12 was associated with a progressive reduction in risk for the total population in primary outcome events and cardiovascular mortality over the 5-year assessment. Although more events occurred in the randomized sibutramine group, on an average, a modest weight loss of approximately 3 kg achieved in the Lead-in Period appeared to offset this increased event rate. Moderate weight loss (3-10 kg) reduced cardiovascular deaths in those with severe, moderate or mild cardiovascular disease. CONCLUSIONS: Modest weight loss over short-term (6 weeks) and longer-term (6-12 months) periods is associated with reduction in subsequent cardiovascular mortality for the following 4-5 years even in those with pre-existing cardiovascular disease. While the sibutramine group experienced more primary outcome events than the placebo group, greater weight loss reduced overall risk of these occurring in both groups.

Weight and blood pressure response to weight management and sibutramine in diabetic and non-diabetic high-risk patients: an analysis from the 6-week lead-in period of the sibutramine cardiovascular outcomes (SCOUT) trial.

OBJECTIVE: To assess treatment responses to sibutramine and weight management in diabetic patients during the lead-in period of the Sibutramine Cardiovascular OUTcomes (SCOUT) trial. METHODS: SCOUT is an ongoing, prospective, randomized, double-blind, placebo-controlled outcome trial in cardiovascular high-risk overweight/obese patients. A total of 10 742 patients received single-blind sibutramine and individualized weight management during the 6-week lead-in period; 84% had a history of type 2 diabetes mellitus and additional co-morbidities. Post-hoc analyses assessed anthropomorphic and vital sign responses between patients with and without diabetes. RESULTS: Concomitant antidiabetic medication use was reported by 86% of the diabetic patients (approximately 30% required insulin-alone or in combination). Body weight and waist circumference decreased in diabetic patients: median 2.1 kg; 2.0 cm (both men and women); for those on insulin: 1.9 kg; 1.5/2.0 cm (men/women); without insulin: 2.3 kg; 2.0 cm (both men and women); blood pressure (BP) was also reduced (median systolic/diastolic 3.5/1.0 mmHg) with larger reductions in diabetic patients who were hypertensive and/or lost the most weight (>5%). In diabetic patients who entered with BP at target (<130/<85 mmHg) but did not lose weight (N = 245), increases of 3.5/2.0 mmHg were observed. Non-diabetic patients had greater weight losses (2.5 kg) but smaller reductions in BP (systolic/diastolic -2.5/-0.5 mmHg). Pulse rate increases were less in diabetic vs. non-diabetic patients (1.5 vs. 2.0 bpm). CONCLUSION: In these high-risk diabetic patients, sibutramine and lifestyle modifications for 6 weeks resulted in small, but clinically relevant, median reductions in body weight, waist circumference and BP. A small median increase in pulse rate was recorded.

Blood pressure changes associated with sibutramine and weight management - an analysis from the 6-week lead-in period of the sibutramine cardiovascular outcomes trial (SCOUT).

OBJECTIVE: To explore vital sign changes among patient subgroups during the 6-week lead-in period of the sibutramine cardiovascular outcomes (SCOUT) trial. METHODS: SCOUT is an ongoing, double-blind, randomized, placebo-controlled outcome trial in overweight/obese patients at high risk of a cardiovascular event. During the 6-week lead-in period, 10,742 patients received sibutramine and weight management. Vital sign changes were assessed post hoc by initial blood pressure (mmHg) categorized as normal (<130/<85), high-normal (130 to <140/85 to <90) or hypertensive (>or=140/>or=90); weight change categories (weight gain/no weight change, >0 to 2.5% weight loss, >2.5 to 5% weight loss and >5% weight loss) and current antihypertensive medication class use (none, one, or two or more). To assess the impact of sibutramine on blood pressure and pulse rate, only patients (N = 10,025) who reported no change in the class of antihypertensive medication used and who did not report an increase in antihypertensive medication use were analysed. RESULTS: At entry, approximately 50% of patients were hypertensive and 26% were high-normal. In hypertensive patients, blood pressure changes (mmHg) decreased by median [5th, 95th percentile] of -6.5 systolic [-27.0, 8.0] and -2.0 diastolic [-15.0, 8.0] (p < 0.001). Hypertensive patients with no weight loss or with weight gain had median decreases of -3.5 systolic [-26.0, 10.0] and -1.5 diastolic [-16.0, 9.0] (p < 0.001). Normotensive patients had median increases of 1.5 systolic [-15.0, 19.5] and 1.0 diastolic [-10.5, 13.0] (p < 0.001) attenuated with increasing weight loss. Approximately 43% of patients initially categorized as hypertensive had a lower blood pressure category at end-point. Concomitant antihypertensive medication classes did not affect blood pressure reductions. Pulse rates were uniformly elevated (median 1-4 bpm, p < 0.001) across blood pressure and weight change categories. CONCLUSIONS: In hypertensive patients (>or=140/>or=90), blood pressure decreases were observed during 6-week treatment with sibutramine even when body weight was unchanged. In patients with normal blood pressure (<130/<85), weight loss of >5% induced decreases in systolic blood pressure; otherwise, small increases were observed. Small pulse rate increases were observed regardless of blood pressure or weight change status.

Olfactory discrimination ability of CD-1 mice for a large array of enantiomers.

With use of a conditioning paradigm, the ability of eight CD-1 mice to distinguish between 15 enantiomeric odor pairs was investigated. The results demonstrate a) that CD-1 mice are capable of discriminating between all odor pairs tested, b) that the enantiomeric odor pairs clearly differed in their degree of discriminability and thus in their perceptual similarity, and c) that pre-training with the rewarded stimuli led to improved initial but not terminal or overall performance. A comparison between the proportion of discriminated enantiomeric odor pairs of the CD-1 mice and those of other species tested in earlier studies on the same discrimination tasks (or on subsets thereof) shows a significant positive correlation between discrimination performance and the number of functional olfactory receptor genes. These findings provide the first evidence of a highly developed ability of CD-1 mice to discriminate between an array of non-pheromonal chiral odorants. Further, they suggest that a species' olfactory discrimination capabilities for these odorants may be correlated with its number of functional olfactory receptor genes. The data presented here may provide useful information for the interpretation of findings from electrophysiological or imaging studies in the mouse and the elucidation of odor structure-activity relationships.

The olfactory receptor family album.

Analysis of the human genome draft sequences has revealed a more complete portrait of the olfactory receptor gene repertoire in humans than was available previously. The new information provides a basis for deeper analysis of the functions of the receptors, and promises new insights into the evolutionary history of the family.

Functional mosaic organization of mouse olfactory receptor neurons.

In contrast to rapid progress in the molecular biology of olfaction, there are few physiological data to characterize the odor response properties of different populations of olfactory receptor neurons (ORNs) and their spatial distributions across the epithelium, which is essential for understanding the coding mechanisms underlying odor discrimination and recognition. We have tested the hypothesis that the ORNs are arranged in a functional mosaic, using an intact epithelial preparation from the mouse, in which odor responses of many ORNs in situ can be monitored simultaneously with calcium imaging techniques. ORNs responding to a given odor were widely distributed across epithelium and intermingled with ORNs responding to other odors. Tight clusters of ORNs responding to the same odor were observed. For a given odor, more ORNs were recruited when the concentration was increased. ORNs were able to distinguish between pairs of enantiomers by showing distinct but somewhat overlapping patterns. The results provide evidence regarding the response spectra of ORNs in situ, supporting the combinatorial coding of odor quality and intensity by different ORN subsets.

Molecular mechanisms underlying differential odor responses of a mouse olfactory receptor.

The prevailing paradigm for G protein-coupled receptors is that each receptor is narrowly tuned to its ligand and closely related agonists. An outstanding problem is whether this paradigm applies to olfactory receptor (ORs), which is the largest gene family in the genome, in which each of 1,000 different G protein-coupled receptors is believed to interact with a range of different odor molecules from the many thousands that comprise "odor space." Insights into how these interactions occur are essential for understanding the sense of smell. Key questions are: (i) Is there a binding pocket? (ii) Which amino acid residues in the binding pocket contribute to peak affinities? (iii) How do affinities change with changes in agonist structure? To approach these questions, we have combined single-cell PCR results [Malnic, B., Hirono, J., Sato, T. & Buck, L. B. (1999) Cell 96, 713-723] and well-established molecular dynamics methods to model the structure of a specific OR (OR S25) and its interactions with 24 odor compounds. This receptor structure not only points to a likely odor-binding site but also independently predicts the two compounds that experimentally best activate OR S25. The results provide a mechanistic model for olfactory transduction at the molecular level and show how the basic G protein-coupled receptor template is adapted for encoding the enormous odor space. This combined approach can significantly enhance the identification of ligands for the many members of the OR family and also may shed light on other protein families that exhibit broad specificities, such as chemokine receptors and P450 oxidases.

Analysis of relations between NMDA receptors and GABA release at olfactory bulb reciprocal synapses.

In the mammalian olfactory bulb, signal processing is mediated by synaptic interactions between dendrites. Glutamate released from mitral cell dendrites excites dendritic spines of granule cells, which in turn release GABA back onto the mitral cell dendrites, forming a reciprocal synaptic pair. This feedback synaptic circuit was shown to be mediated predominantly by NMDA receptors. We further utilized caged Ca2+ compounds to obtain insight into the mechanism that couples NMDA receptor activation to GABA release. Feedback inhibition elicited by photo-release of caged Ca2+ in mitral cell secondary dendrites persisted when voltage-gated Ca2+ channels were blocked by cadmium (Cd2+) and nickel (Ni2+). These results indicate that Ca2+ influx through NMDA receptors can directly trigger presynaptic GABA release for local dendrodendritic feedback inhibition.

Olfactory receptor database: a sensory chemoreceptor resource.

The Olfactory Receptor Database (ORDB) is a WWW-accessible database that has been expanded from an olfactory receptor resource to a chemoreceptor resource. It stores data on six classes of G-protein-coupled sensory chemoreceptors: (i) olfactory receptor-like proteins, (ii) vomeronasal receptors, (iii) insect olfactory receptors, (iv) worm chemo-receptors, (v) taste papilla receptors and (vi) fungal pheromone receptors. A complementary database of the ligands of these receptors (OdorDB) has been constructed and is publicly available in a pilot mode. The database schema of ORDB has been changed from traditional relational to EAV/CR (Entity-Attribute-Value with Classes and Relationships), which allows the interoperability of ORDB with other related databases as well as the creation of intra-database associations among objects. This inter-operability facilitates users to follow information from odor molecule binding to its putative receptor, to the properties of the neuron expressing the receptor, to a computational model of activity of olfactory bulb neurons. In addition, tools and resources have been added allowing users to access interactive phylogenetic trees and alignments of sensory chemoreceptors. ORDB is available via the WWW at http://ycmi.med. yale.edu/senselab/ordb/

Electrophysiological characterization of rat and mouse olfactory receptor neurons from an intact epithelial preparation.

To understand the coding mechanisms underlying olfactory discrimination, it is necessary to characterize odor response properties of olfactory receptor neurons (ORNs). In contrast with rapid progress in molecular biology, there is little physiological data from ORNs in rodent. To facilitate acquisition of such data, we have developed an intact olfactory epithelial preparation from both rat and mouse. We have carried out initial studies of this preparation by monitoring odor responses by patch-clamping directly on the ORN dendritic knobs, a subcellular site very close to the locus of olfactory signal transduction. Our results show that rat and mouse ORNs have similar intrinsic membrane properties. Most cells fired spontaneously at a low frequency (f) and about one half fired repetitively in response to current (I) injection with a linear f/I relation. About one third of rat and mouse ORNs responded to a mixture of four odors in a dose-dependent manner and about 60% of them responded to IBMX, a potent inhibitor of phosphodiesterase. The results suggest that this intact preparation offers the advantage of approximating in vivo physiological conditions, while furnishing an opportunity to map single neuron responses in the epithelium in a spatially-defined manner, using electrophysiological or cell imaging methods.

Computational analysis of action potential initiation in mitral cell soma and dendrites based on dual patch recordings.

In olfactory mitral cells, dual patch recordings show that the site of action potential initiation can shift between soma and distal primary dendrite and that the shift is dependent on the location and strength of electrode current injection. We have analyzed the mechanisms underlying this shift, using a model of the mitral cell that takes advantage of the constraints available from the two recording sites. Starting with homogeneous Hodgkin-Huxley-like Na(+)-K(+) channel distribution in the soma-dendritic region and much higher sodium channel density in the axonal region, the model's channel kinetics and density were adjusted by a fitting algorithm so that the model response was virtually identical to the experimental data. The combination of loading effects and much higher sodium channel density in the axon relative to the soma-dendritic region results in significantly lower "voltage threshold" for action potential initiation in the axon; the axon therefore fires first unless the voltage gradient in the primary dendrite is steep enough for it to reach its higher threshold. The results thus provide a quantitative explanation for the stimulus strength and position dependence of the site of action potential initiation in the mitral cell.

Synaptic organization and neurotransmitters in the rat accessory olfactory bulb.

The accessory olfactory bulb (AOB) is the first relay station in the vomeronasal system and may play a critical role in processing pheromone signals. The AOB shows similar but less distinct lamination compared with the main olfactory bulb (MOB). In this study, synaptic organization of the AOB was analyzed in slice preparations from adult rats by using both field potential and patch-clamp recordings. Stimulation of the vomeronasal nerve (VN) evoked field potentials that showed characteristic patterns in different layers of the AOB. Current source density (CSD) analysis of the field potentials revealed spatiotemporally separated loci of inward current (sinks) that represented sequential activation of different neuronal components: VN activity (period I), synaptic excitation of mitral cell apical dendrites (period II), and activation of granule cells by mitral cell basal dendrites (period III). Stimulation of the lateral olfactory tract also evoked field potentials in the AOB, which indicated antidromic activation of the mitral cells (period I and II) followed by activation of granule cells (period III). Whole cell patch recordings from mitral and granule cells of the AOB supported that mitral cells are excited by VN terminals and subsequently activate granule cells through dendrodendritic synapses. Both CSD analysis and patch recordings provided evidence that glutamate is the neurotransmitter at the vomeronasal receptor neuron; mitral cell synapses and both NMDA and non-NMDA receptors are involved. We also demonstrated electrophysiologically that reciprocal interaction between mitral and granule cells in the AOB is through the dendrodendritic reciprocal synapses. The neurotransmitter at the mitral-to-granule synapses is glutamate and at the granule-to-mitral synapse is gamma-aminobutyric acid. The synaptic interactions among receptor cell terminals, mitral cells, and granule cells in the AOB are therefore similar to those in the MOB, suggesting that processing of chemosensory information in the AOB shares similarities with that in the MOB.

Olfactory Receptor Database: a database of the largest eukaryotic gene family.

The Olfactory Receptor Database (ORDB) is a WWW-accessible database that stores data on Olfactory Receptor-like molecules (ORs) and has been open to the public since June 1996. It contains a public and a private area. The public area includes published DNA and protein sequence data for ORs, links to OR models and data on their expression, chromosomal localization and source organism, as well as (i) links to bibliography through PubMed and (ii) interactive WWW-based tools, such as BLAST homology searching. The private area functions as a service to laboratories that are actively cloning receptors. Source laboratories enter the sequences of the receptor clones they have characterized to the private database and can search for identical or near identical OR sequences in both public and private databases. If another laboratory has cloned and deposited an identical or closely matching sequence there are means for communication between the laboratories to help avoid duplication of work. ORDB is available via the WWW at http://crepe.med.yale.edu/ORDB/HTML

Identification of olfactory receptor mRNA sequences from the rat olfactory bulb glomerular layer.

In situ hybridization has demonstrated mRNA for olfactory receptors (OR) in the axon terminals of olfactory receptor neurons. Neurons that express the same OR appear to send their axons to two stereotyped glomeruli in the olfactory bulb (OB). Based on these observations, we tested the feasibility of using RT-PCR to isolate and sequence OR mRNA from small samples of the rat OB glomerular layer. Biomagnetic mRNA isolation followed by RT-PCR yielded partial sequences for 21 novel members of the OR family. The results suggest that the topography of OR mRNA can be mapped across the OB, to study synaptic specificity and odor representation in the olfactory system.

The Human Brain Project: neuroinformatics tools for integrating, searching and modeling multidisciplinary neuroscience data.

What is neuroinformatics? What is the Human Brain Project? Why should you care? Supported by a consortium of US funding agencies, the Human Brain Project aims to bring to the analysis of brain function the same advantages of Internet-accessible databases and database tools that have been crucial to the development of molecular biology and the Human Genome Project. The much greater complexity of neural data, however, makes this a far more challenging task. As a pilot project in this new initiative, we review some of the progress that has been made and indicate some of the problems, challenges and opportunities that lie ahead.

Three-dimensional structure and composition of CA3-->CA1 axons in rat hippocampal slices: implications for presynaptic connectivity and compartmentalization.

Physiological studies of CA3-->CA1 synaptic transmission and plasticity have revealed both pre- and postsynaptic effects. Understanding the extent to which individual presynaptic axonal boutons could provide local compartments for control of synaptic efficacy and microconnectivity requires knowledge of their three-dimensional morphology and composition. In hippocampal slices, serial electron microscopy was used to examine a nearly homogeneous population of CA3-->CA1 axons in the middle of stratum radiatum of area CA1. The locations of postsynaptic densities (PSDs), vesicles, and mitochondria were determined along 75 axon segments (9.1 +/- 2.0 micrometer in length). Synapses, defined by the colocalization of PSDs and vesicles, occurred on average at 2.7 micrometer intervals along the axons. Most varicosities (68%) had one PSD, 19% had 2-4 PSDs, and 13% had none. Synaptic vesicles occurred in 90% of the varicosities. One-half (53%) of the varicosities lacked mitochondria, raising questions about their regulation of ATP and Ca2+, and 8% of varicosities contained only mitochondria. Eleven axons were reconstructed fully. The varicosities were oblong and varied greatly in both length (1.1 +/- 0.7 micrometer) and volume (0.13 +/- 0.14 micrometer 3), whereas the intervaricosity shafts were narrow, tubular, and similar in diameter (0.17 +/- 0.04 micrometer) but variable in length (1.4 +/- 1.2 micrometer). The narrow axonal shafts resemble dendritic spine necks and thus could promote biochemical compartmentalization of individual axonal varicosities. The findings raise the intriguing possibility of localized differences in metabolism and connectivity among different axons, varicosities, and synapses.

Imaging odor-induced calcium transients in single olfactory cilia: specificity of activation and role in transduction.

The possibility that odor stimuli trigger distinct Ca2+ elevations within the cilia of vertebrate olfactory receptor neurons (ORNs) is a widely proposed concept. However, because of the small size of the olfactory cilia, the existence and properties of such Ca2+ elevations and their role in odor transduction are still unknown. We investigate odor-induced Ca2+ changes in individual olfactory cilia from salamander using the Ca2+ indicator dye fluo-3 in combination with laser scanning confocal microscopy. Single brief applications of odor ligand produce highly localized Ca2+ elevations in individual cilia lasting for several seconds. These Ca2+ signals originate in the cilia and depend entirely on Ca2+ entry through ciliary cyclic nucleotide-gated ion channels. The odor specificity of the Ca2+ rises implies a receptor-operated mechanism underlying odor detection. Each of the cilia on a receptor neuron functions as an independent biochemical compartment that can detect odorants and produce a Ca2+ transient with remarkably uniform properties in terms of kinetics and odor specificity. The rate of recovery of the odor-induced Ca2+ transients matches recovery from a short-term form of odor adaptation. Application of the membrane-permeant intracellular Ca2+ chelator BAPTA AM eliminates this odor adaptation. The results indicate that an olfactory cilium serves as a basic functional unit at the input level of the olfactory system, controlling both the specificity and sensitivity of odor detection.