Comparison of the monoamine transporters from human and mouse in their sensitivities to psychostimulant drugs

BACKGROUND: The plasma membrane neurotransmitter transporters terminate neurotransmissions by the reuptake of the released neurotransmitters. The transporters for the monoamines dopamine, norepinephrine, and serotonin (DAT, NET, and SERT) are targets for several popular psychostimulant drugs of abuse. The potencies of the psychostimulant on the monoamine transporters have been studied by several laboratories. However, there are significant discrepancies in the reported data with differences up to 60-fold. In addition, the drug potencies of the 3 monoamine transporters from mouse have not been compared in the same experiments or along side the human transporters. Further studies and systematic comparisons are needed. RESULTS: In this study, we compared the potencies of five psychostimulant drugs to inhibit human and mouse DAT, SERT and NET in the same cellular background. The K(I )values of cocaine to inhibit the 3 transporters are within a narrow range of 0.2 to 0.7 μM. In comparison, methylphenidate inhibited DAT and NET at around 0.1 μM, while it inhibited SERT at around 100 μM. The order of amphetamine potencies was NET (K(I )= 0.07–0.1 μM), DAT (K(I )≈ 0.6 μM), and SERT (K(I )between 20 to 40 μM). The results for methamphetamine were similar to those for amphetamine. In contrast, another amphetamine derivative, MDMA (3–4 methylenedioxymethamphetamine), exhibited higher potency at SERT than at DAT. The human and mouse transporters were similar in their sensitivities to each of the tested drugs (K(I )values are within 4-fold). CONCLUSION: The current and previous studies support the following conclusions: 1) cocaine blocks all 3 monoamine transporters at similar concentrations; 2) methylphenidate inhibits DAT and NET well but a 1000-fold higher concentration of the drug is required to inhibit SERT; 3) Amphetamine and methamphetamine are most potent at NET, while being 5- to 9-fold less potent at DAT, and 200- to 500-fold less potent at SERT; 4) MDMA has moderately higher apparent affinity for SERT and NET than for DAT. The relative potencies of a drug to inhibit DAT, NET and SERT suggest which neurotransmitter systems are disrupted the most by each of these stimulants and thus the likely primary mechanism of drug action

Cocaine reward and locomotion stimulation in mice with reduced dopamine transporter expression

<p>Abstract</p> <p>Background</p> <p>The dopamine transporter (DAT) plays a critical role in regulating dopamine neurotransmission. Variations in DAT or changes in basal dopaminergic tone have been shown to alter behavior and drug responses. DAT is one of the three known high affinity targets for cocaine, a powerful psychostimulant that produces reward and stimulates locomotor activity in humans and animals. We have shown that cocaine no longer produces reward in knock-in mice with a cocaine insensitive mutant DAT (DAT-CI), suggesting that cocaine inhibition of DAT is critical for its rewarding effect. However, in DAT-CI mice, the mutant DAT has significantly reduced uptake activity resulting in elevated basal dopaminergic tone, which might cause adaptive changes that alter responses to cocaine. Therefore, the objective of this study is to determine how elevated dopaminergic tone affects how mice respond to cocaine.</p> <p>Results</p> <p>We examined the cocaine induced behavior of DAT knockdown mice that have DAT expression reduced by 90% when compared to the wild type mice. Despite a dramatic reduction of DAT expression and marked elevation in basal dopamine tone, cocaine produced reward, as measured by conditioned place preference, and stimulated locomotor activity in these mice.</p> <p>Conclusion</p> <p>A reduction in DAT expression and elevation of dopaminergic tone do not lead to adaptive changes that abolish the rewarding and stimulating effects of cocaine. Therefore, the lack of reward to cocaine observed in DAT-CI mice is unlikely to have resulted from the reduced DAT activity but instead is likely due to the inability of cocaine to block the mutated DAT and increase extracellular dopamine. This study supports the conclusion that the blockade of DAT is required for cocaine reward and locomotor stimulation.</p

Potencies of Cocaine Methiodide on Major Cocaine Targets in Mice

Cocaine methiodide (CM), a charged cocaine analog, cannot pass the blood brain barrier. It has been assumed the effects of systemic CM represent cocaine actions in peripheral tissues. However, the IC50 values of CM have not been clearly determined for the major cocaine targets: dopamine, norepinephrine, and serotonin transporters, and sodium channels. Using cells transfected with individual transporters from mice and synaptosomes from mouse striatum tissues, we observed that the inhibition IC50 values for monoamine uptake by CM were 31-fold to 184-fold higher compared to cocaine at each of the transporters. In dorsal root ganglion neurons, cocaine inhibited sodium channels with an apparent IC50 of 75 µM, while CM showed no observable effect at concentrations up to 3 mM. These results indicate that an equal dose of CM will not produce an equivalent peripheral effect of cocaine

Bulk Cutting of Carbon Nanotubes Using Electron Beam Irradiation

According to some embodiments, the present invention provides a method for attaining short carbon nanotubes utilizing electron beam irradiation, for example, of a carbon nanotube sample. The sample may be pretreated, for example by oxonation. The pretreatment may introduce defects to the sidewalls of the nanotubes. The method is shown to produces nanotubes with a distribution of lengths, with the majority of lengths shorter than 100 tun. Further, the median length of the nanotubes is between about 20 nm and about 100 nm

A Bioequivalence Test by the Direct Comparison of Concentration-versus-Time Curves Using Local Polynomial Smoothers

In order to test if two chemically or pharmaceutically equivalent products have the same efficacy and/or toxicity, a bioequivalence (BE) study is conducted. The 80%/125% rule is the most commonly used criteria for BE and states that BE cannot be claimed unless the 90% CIs for the ratio of selected pharmacokinetics (PK) parameters of the tested to the reference drug are within 0.8 to 1.25. Considering that estimates of these PK parameters are derived from the concentration-versus-time curves, a direct comparison between these curves motivates an alternative and more flexible approach to test BE. Here, we propose to frame the BE test in terms of an equivalence of concentration-versus-time curves which are constructed using local polynomial smoother (LPS). A metric is presented to quantify the distance between the curves and its 90% CIs are calculated via bootstrapping. Then, we applied the proposed procedures to data from an animal study and found that BE between a generic drug and its brand name cannot be concluded, which was consistent with the results by applying the 80%/125% rule. However, the proposed procedure has the advantage of testing only on a single metric, instead of all PK parameters

Foot Loading Characteristics of Chinese Bound Feet Women: A Comparative Analysis

The custom of bound feet among Chinese women has existed for almost a century. This practice has influenced the daily life of Chinese women, especially during everyday locomotion. The primary aim of this study is to analyze the loading patterns of bound feet. Specifically, the plantar pressure and center of pressure were analyzed for peak pressure, contact area, force time integral, center of pressure displacement velocity and trajectory in the anterior- posterior direction via a comparison with normal feet. The key outcomes from this work were that the forefoot and rearfoot of bound feet bear the whole loading during stance phase. The center of pressure displacement velocity of bound feet was also greatly reduced with the shortening of trajectories. This suggests that the proprioceptive system adjusts motor function to adapt to new loading patterns while maintaining locomotive stability. A biomechanical understanding of bound feet may assist with prevention, treatment and rehabilitation of bound feet disorders

Impact of Community-Based Larviciding on the Prevalence of Malaria Infection in Dar es Salaam, Tanzania.

The use of larval source management is not prioritized by contemporary malaria control programs in sub-Saharan Africa despite historical success. Larviciding, in particular, could be effective in urban areas where transmission is focal and accessibility to Anopheles breeding habitats is generally easier than in rural settings. The objective of this study is to assess the effectiveness of a community-based microbial larviciding intervention to reduce the prevalence of malaria infection in Dar es Salaam, United Republic of Tanzania. Larviciding was implemented in 3 out of 15 targeted wards of Dar es Salaam in 2006 after two years of baseline data collection. This intervention was subsequently scaled up to 9 wards a year later, and to all 15 targeted wards in 2008. Continuous randomized cluster sampling of malaria prevalence and socio-demographic characteristics was carried out during 6 survey rounds (2004-2008), which included both cross-sectional and longitudinal data (N = 64,537). Bayesian random effects logistic regression models were used to quantify the effect of the intervention on malaria prevalence at the individual level. Effect size estimates suggest a significant protective effect of the larviciding intervention. After adjustment for confounders, the odds of individuals living in areas treated with larviciding being infected with malaria were 21% lower (Odds Ratio = 0.79; 95% Credible Intervals: 0.66-0.93) than those who lived in areas not treated. The larviciding intervention was most effective during dry seasons and had synergistic effects with other protective measures such as use of insecticide-treated bed nets and house proofing (i.e., complete ceiling or window screens). A large-scale community-based larviciding intervention significantly reduced the prevalence of malaria infection in urban Dar es Salaam

Evaluation of commercially available RNA amplification kits for RNA sequencing using very low input amounts of total RNA

This article includes supplemental data. Please visit http://www.fasebj.org to obtain this information.Multiple recent publications on RNA sequencing (RNA-seq) have demonstrated the power of next-generation sequencing technologies in whole-transcriptome analysis. Vendor-specific protocols used for RNA library construction often require at least 100 ng total RNA. However, under certain conditions, much less RNA is available for library construction. In these cases, effective transcriptome profiling requires amplification of subnanogram amounts of RNA. Several commercial RNA amplification kits are available for amplification prior to library construction for next-generation sequencing, but these kits have not been comprehensively field evaluated for accuracy and performance of RNA-seq for picogram amounts of RNA. To address this, 4 types of amplification kits were tested with 3 different concentrations, from 5 ng to 50 pg, of a commercially available RNA. Kits were tested at multiple sites to assess reproducibility and ease of use. The human total reference RNA used was spiked with a control pool of RNA molecules in order to further evaluate quantitative recovery of input material. Additional control data sets were generated from libraries constructed following polyA selection or ribosomal depletion using established kits and protocols. cDNA was collected from the different sites, and libraries were synthesized at a single site using established protocols. Sequencing runs were carried out on the Illumina platform. Numerous metrics were compared among the kits and dilutions used. Overall, no single kit appeared to meet all the challenges of small input material. However, it is encouraging that excellent data can be recovered with even the 50 pg input total RNA