DAT isn't all that: cocaine reward and reinforcement require Toll-like receptor 4 signaling.

The initial reinforcing properties of drugs of abuse, such as cocaine, are largely attributed to their ability to activate the mesolimbic dopamine system. Resulting increases in extracellular dopamine in the nucleus accumbens (NAc) are traditionally thought to result from cocaine's ability to block dopamine transporters (DATs). Here we demonstrate that cocaine also interacts with the immunosurveillance receptor complex, Toll-like receptor 4 (TLR4), on microglial cells to initiate central innate immune signaling. Disruption of cocaine signaling at TLR4 suppresses cocaine-induced extracellular dopamine in the NAc, as well as cocaine conditioned place preference and cocaine self-administration. These results provide a novel understanding of the neurobiological mechanisms underlying cocaine reward/reinforcement that includes a critical role for central immune signaling, and offer a new target for medication development for cocaine abuse treatment.

Synthesis of indoles, benzofurans, and related heterocycles via an acetylene-activated SNAr/intramolecular cyclization cascade sequence in water or DMSO.

The synthesis of 2-substituted indoles and benzofurans was achieved by nucleophilic aromatic substitution, followed by subsequent 5-endo-dig cyclization between the nucleophile and an ortho acetylene. The acetylene serves the dual role of the electron withdrawing group to activate the substrate for SNAr, and the C1-C2 carbon scaffold for the newly formed 5-membered heteroaromatic ring. This method allows for the bond forming sequence of Ar-X-N/O-C1 to proceed in a single synthetic step, furnishing indoles and benzofurans in moderate to high yields. Since the method is not transition metal mediated, brominated and chlorinated substrates are tolerated, and benzofuran formation can be conducted using water or water-DMSO mixtures as solvent.

Opioid activation of toll-like receptor 4 contributes to drug reinforcement.

Opioid action was thought to exert reinforcing effects solely via the initial agonism of opioid receptors. Here, we present evidence for an additional novel contributor to opioid reward: the innate immune pattern-recognition receptor, toll-like receptor 4 (TLR4), and its MyD88-dependent signaling. Blockade of TLR4/MD2 by administration of the nonopioid, unnatural isomer of naloxone, (+)-naloxone (rats), or two independent genetic knock-outs of MyD88-TLR4-dependent signaling (mice), suppressed opioid-induced conditioned place preference. (+)-Naloxone also reduced opioid (remifentanil) self-administration (rats), another commonly used behavioral measure of drug reward. Moreover, pharmacological blockade of morphine-TLR4/MD2 activity potently reduced morphine-induced elevations of extracellular dopamine in rat nucleus accumbens, a region critical for opioid reinforcement. Importantly, opioid-TLR4 actions are not a unidirectional influence on opioid pharmacodynamics, since TLR4(-/-) mice had reduced oxycodone-induced p38 and JNK phosphorylation, while displaying potentiated analgesia. Similar to our recent reports of morphine-TLR4/MD2 binding, here we provide a combination of in silico and biophysical data to support (+)-naloxone and remifentanil binding to TLR4/MD2. Collectively, these data indicate that the actions of opioids at classical opioid receptors, together with their newly identified TLR4/MD2 actions, affect the mesolimbic dopamine system that amplifies opioid-induced elevations in extracellular dopamine levels, therefore possibly explaining altered opioid reward behaviors. Thus, the discovery of TLR4/MD2 recognition of opioids as foreign xenobiotic substances adds to the existing hypothesized neuronal reinforcement mechanisms, identifies a new drug target in TLR4/MD2 for the treatment of addictions, and provides further evidence supporting a role for central proinflammatory immune signaling in drug reward.

Photoinduced nucleation of water vapor.

Strong photoinduced nucleation of pure water vapor was found to occur in a wavelength range where no ultraviolet absorption of water vapor has been reported. Systematic studies were made of the dependence of the nucleation rate and the delay time for the initiation of nucleation on light intensity. The results obtained were accurately fitted by a phenomenological mechanism whereby the nucleation is initiated by clusters accumulating an appropriate number of photoexcited water molecules.

Slow compressional wave propagation in wet human and bovine cortical bone.

Ultrasonic wave propagation in bovine plexiform and human Haversian bone was studied in the range 0.5 to 15 megahertz. A new longitudinal wave was observed which traveled more slowly than the ordinary longitudinal wave. The slow wave was associated with the dynamics of fluid motion in the pores of bone.

Nucleation on photoexcited molecules.

On irradiation with light of suitable wavelength and intensity, certain organic compounds, even at very low concentrations, cause very efficient nucleation of supersaturated vapors. A mechanism is suggested to account for this phenomenon. Nuclei containing only a few photoexcited molecules are responsible for the nucleation.

Poly(meta-phenylene oxides) for the design of a tunable, efficient, and reusable catalytic platform.

We present poly(meta-phenylene oxides) as versatile and tunable scaffolds for immobilized catalyst design. Their modular synthesis enables the rational incorporation of different side chain groups with positional control along the polymer backbone, providing an easy means to modulate polymer properties, and their corresponding catalytic activity. The coordinative convolution of these polymers with (NH4)2PdCl4 affords efficient, reusable catalysts for various organic transformations.

Anorexia nervosa and gonadal dysgenesis. Further evidence of a relationship.

We report the ninth documentated coincidence of anorexia nervosa and XO gonadel dysgenesis (Turner syndrome) and the second confirmed coincidence of anorexia nervosa and XO/XX (mosaic) gonadal dysgenesis. Three other women who had anorexia nervosa and clinical stigmata of gonadal dysgenesis, but who were not karyotyped, are also described. We thus consider the possibility that patients with gonadal dysgenesis are at special risk for anorexia nervosa and discuss hypotheses to explain such a relationship.

The production rate of cortisol declines during recovery from anorexia nervosa.

The plasma concentration of cortisol is elevated in many patients with anorexia nervosa. It has remained unclear whether this elevation of plasma cortisol level is due only to a slowing of the rate of cortisol metabolism or whether there is, as well, an increase in adrenal secretory activity in anorexia nervosa. We studied adrenocortical activity in 9 female patients and one male patient with anorexia nervosa before and during recovery. The 24-h mean level of plasma cortisol and the rate of urinary free cortisol excretion decreased during recovery, from 11.4 to 7.4 micrograms/dl and from 225 to 116 micrograms/day, respectively (P less than 0.005 and P less than 0.10, paired t test). These changes were associated with a significant decline in the rate of cortisol production from 24.3 to 17.9 mg/day as measured by radioisotope dilution (P less than 0.005). These results suggest that adrenal secretory activity is increased in anorexia nervosa and that the elevation of plasma cortisol level observed in this syndrome reflects not only a slowing of cortisol metabolism but also a rise in cortisol production.

Weaning and depression: another postpartum complication.

The dramatic endocrinologic changes that occur after childbirth and the coincident affective disturbances that range from "maternity blues" to major depression have stimulated much theorizing and some study. The role of breast-feeding and weaning has received remarkably little attention both in the more biologically oriented studies and in epidemiologic work. This paper reviews endocrinologic data which support the thesis that postpartum psychiatric disorders have a hormonal basis and discusses the possible psychiatric effects of breast-feeding and weaning. The cases of four patients who developed major depressions in close temporal association with weaning are presented and discussed.

Is there a relationship between eating disorder and affective disorder? New evidence from sleep recordings.

There is evidence that patients with anorexia nervosa (particularly those who also have bulimia) and patients with affective disorder share many features. The authors present sleep polygraph data from 20 young women with anorexia nervosa (17 also bulimic) and 10 age-matched normal women. Their urinary free cortisol levels were determined, and the subjects with eating disorders were also rated for depression. The findings suggest the existence of a subgroup of patients who show sleep abnormalities, in addition to clinical and possibly endocrine abnormalities, that indicate concurrent affective disorder. The authors present several models that could account for this relationship.

Cerebral glucose metabolism in women with panic disorder.

OBJECTIVE: This study was undertaken to clarify earlier inconsistent findings in brain metabolic topography in panic disorder patients at rest. METHOD: Positron emission tomography (PET) with [18F]fluorodeoxyglucose was used to determine cerebral metabolic activity in six female patients with a DSM-III-R diagnosis of panic disorder and in six healthy female volunteers. All patients with panic disorder were medication free and were sensitive to lactate infusion. RESULTS: A significant increase in glucose metabolism was found in the left hippocampus and parahippocampal area of the panic disorder subjects in comparison with that found in the healthy subjects. In addition, a significant decrease in metabolism was found in the right inferior parietal and right superior temporal brain regions of the panic disorder subjects in comparison with that of the normal subjects. There was no significant correlation between scores for the severity of panic disorder or for the severity of lactate-induced panic attack and the quantified PET abnormality. CONCLUSIONS: These data provide further support for the hypothesis of an abnormal brain metabolism in the hippocampal and parahippocampal area in individuals with panic disorder and also suggest other areas of aberrant brain metabolism in this disorder.

DNA interstrand cross-linking by a mycotoxic diepoxide.

The diepoxide mycotoxin (2R, 3R, 8R, 9R)-4,6-decadiyne-2,3:8,9-diepoxy-1,10-diol (repandiol) was both isolated from the mushroom Hydnum repandum and synthesized de novo. Repandiol was found to form interstrand cross-links within a restriction fragment of DNA, linking deoxyguanosines on opposite strands primarily within the 5'-GNC and 5'-GNNC sequences preferred by diepoxyoctane. However, repandiol was a significantly less efficient cross-linker than either of the diepoxyalkanes (diepoxyoctane and diepoxybutane) to which it was compared.

Rate-dependent effects of d-and l-amphetamine on schedule-controlled responding in pigeons and squirrel monkeys.

The effects of d- and l-amphetamine were studied on key-pressing responses of squirrel monkeys maintained under fixed-interval schedules of electric shock presentation, and on key-pecking responses of pigeons maintained under multiple fixed-ratio, fixed-interval schedules of food presentation. Under the fixed-interval schedules, responding followed a pause and occurred at increasing rates as the interval elapsed. Both isomers produced comparable increases in rates of responding under relatively long fixed-interval schedules with small to intermediate doses; maximal effects of the d-isomer were obtained at doses one-half log unit smaller than the doses of the l-isomer. Responding of pigeons maintained under relatively short-fixed-interval schedules was only decreased by either amphetamine isomer. Responding of pigeons maintained under fixed-ratio schedules occurred at the highest rates and was also only decreased by either amphetamine isomer. Decreased in response rate produced by the d-isomer were generally obtained at doses one-half log unit smaller than doses of the l-isomer than produced comparable effects. Both isomers increased responding that occurred at low rates early in the fixed-interval to a proportionally greater extent than thigh rates from later in the interval. The highest rates in the fixed-interval was generally decreased. Differences in potency between the two isomers in producing rate-dependent effects were small, most noticable with larger doses, and less than the potency differences between the two isomers in producing changes in response rate.

2D QSAR modeling and preliminary database searching for dopamine transporter inhibitors using genetic algorithm variable selection of Molconn Z descriptors.

In light of the chronic problem of abuse of the controlled substance cocaine, we have investigated novel approaches toward both understanding the activity of inhibitors of the dopamine transporter (DAT) and identifying novel inhibitors that may be of therapeutic potential. Our most recent studies toward these ends have made use of two-dimensional (2D) quantitative structure-activity relationship (QSAR) methods in order to develop predictive models that correlate structural features of DAT ligands to their biological activities. Specifically, we have adapted the method of genetic algorithms-partial least squares (GA-PLS) (Cho et al. J. Comput. -Aided Mol. Des., submitted) to the task of variable selection of the descriptors generated by the software Molconn Z. As the successor to the program Molconn X, which generated 462 descriptors, Molconn Z provides 749 chemical descriptors. By employing genetic algorithms in optimizing the inclusion of predictive descriptors, we have successfully developed a robust model of the DAT affinities of 70 structurally diverse DAT ligands. This model, with an exceptional q(2) value of 0.85, is nearly 25% more accurate in predictive value than a comparable model derived from Molconn X-derived descriptors (q(2) = 0.69). Utilizing activity-shuffling validation methods, we have demonstrated the robustness of both this DAT inhibitor model and our QSAR method. Moreover, we have extended this method to the analysis of dopamine D(1) antagonist affinity and serotonin ligand activity, illustrating the significant improvement in q(2) for a variety of data sets. Finally, we have employed our method in performing a search of the National Cancer Institute database based upon activity predictions from our DAT model. We report the preliminary results of this search, which has yielded five compounds suitable for lead development as novel DAT inhibitors.

Novel tropane-based irreversible ligands for the dopamine transporter.

3alpha-(diphenylmethoxy)tropane (benztropine) and its analogues are tropane ring-containing dopamine uptake inhibitors that display binding and behavioral profiles that are distinct from cocaine. We previously prepared a benztropine-based photoaffinity label [125I]-(N-[4-(4'-azido-3'-iodophenyl)butyl]-3alpha-[bis(4'-fluorophenyl)methoxy]tropane, [125I]1, that covalently attached to the 1-2 transmembrane spanning region of the dopamine transporter (DAT). This was in contrast to the 4-7 transmembrane spanning region labeled by a cocaine-based photoaffinity label, [125I] 2 (RTI 82). To characterize further these different binding domains, photoaffinity ligands that had the 4'-azido-3'-iodophenyl substituent extended from the same position on the tropane ring were desirable. Thus, identification of the optimal alkyl linker between this substituent and the tropane nitrogen in the benztropine series was investigated to ultimately prepare the identical N-substituted analogue of 2. In this pursuit, the N-[4-(4'-azido-3'-iodophenyl)propyl] analogue of 3alpha-[bis(4'-fluorophenyl)methoxy]tropane (9a) was synthesized as well as two isothiocyanate analogues that do not require photoactivation (10a,b) for irreversible binding. The synthesis of these target compounds was achieved using a modification of the strategy developed for 1. Evaluation of these compounds for displacing [3H]WIN 35 428 binding at DAT in rat caudate putamen revealed that the 4'-azido-3'-iodophenylbutyl substituent, found in 1, provided optimal binding affinity and was chosen to replace the N-CH3 group on 2. Both the 4'-azido-3'-iodophenyl- and the 4'-isothiocyanatophenylbutyl analogues of 2 (25 and 26, respectively) were synthesized. Both products bound to DAT with comparable potency (IC(50) = 30 nM) to RTI 82 (2). In addition, compound 26 demonstrated wash-resistant displacement of [3H]WIN 35 428 in HEK 293 cells stably transfected with hDAT. These ligands will provide important tools for further characterizing the binding domains for tropane-based dopamine uptake inhibitors at the DAT.

Novel 4'-substituted and 4',4"-disubstituted 3 alpha-(diphenylmethoxy)tropane analogs as potent and selective dopamine uptake inhibitors.

A series of 4'-substituted and 4',4"-disubstituted 3 alpha-(diphenylmethoxy)tropane analogs were prepared as novel probes for the dopamine transporter. These compounds were evaluated in radiolabeled binding assays for the dopamine, norepinephrine, and serotonin transporters. All of these compounds monophasically displaced [3H]WIN 35,428 binding in rat caudate putamen with Ki values ranging from 11.8 to 2000 nM. The most potent compound in this series was 4',4"-difluoro 3 alpha-(diphenylmethoxy)tropane 7c with a Ki = 11.8 nM. All of the compounds inhibited dopamine uptake in rat caudate putamen (IC50 = 24-4456 nM) which correlated significantly (r = 0.907; p > 0.0001) with binding affinities at the dopamine transporter. None of the compounds demonstrated high-affinity binding at the norepinephrine (Ki > 4800 nM) or serotonin (Ki > 690 nM) transporters. Therefore, the most potent dopamine uptake inhibitors in this series were highly selective for the dopamine transporter. Preliminary behavioral studies of several of these analogs (7a-e) suggested that the compounds did not display a cocaine-like behavioral profile, despite their ability to inhibit dopamine uptake. The present data coupled with the observed differences from cocaine in structure-activity relationships suggested that the 3 alpha-(diphenylmethoxy)tropane analogs may be interacting at a different active site than cocaine on the dopamine transporter and that an additional binding domain might be exploited for the identification of potential therapeutics for the treatment of cocaine abuse.

Synthesis and dopamine transporter affinity of 2-(methoxycarbonyl)-9-methyl-3-phenyl-9-azabicyclo[3.3.1]nonane derivatives.

A series of 9-methyl-3 beta-phenyl-2-substituted-9-azabicyclo[3.3.1]nonane derivatives were synthesized and evaluated as cocaine-binding site ligands at the dopamine transporter (DAT). The conformation of the bicyclic structures and the stereochemistry of the substituents were determined by NMR and X-ray crystallography. The in vitro binding affinity (Ki) of the 9-azabicyclo[3.3.1]nonane derivatives was measured in rat caudate-putamen tissue, and they were found to be 100-fold (Ki = 2-14 microM) less potent than cocaine and other tropane analogs. From these results it is evident that the cocaine-binding site at the DAT is very sensitive to structural modifications of the unsubstituted methylene bridge [C(6)-C(7)] of cocaine and cocaine-like compounds.

Synthesis and dopamine transporter affinity of the four stereoisomers of (+/-)-2-(methoxycarbonyl)-7-methyl-3-phenyl-7-azabicyclo[2.2.1]heptane.

All four stereoisomers of (+/-)-2-(methoxycarbonyl)-7-methyl-3-phenyl-7-azabicyclo[2.2. 1]heptane were synthesized and evaluated as cocaine binding site ligands at the dopamine transporter. The in vitro binding affinities (Ki) of the 7-azabicyclo[2.2.1]heptane derivatives were measured in rat caudate-putamen tissue and found to be 100-3000-fold less potent (Ki = 5-96 microM) than cocaine and 2beta-(methoxycarbonyl)-3beta-phenyltropane (2, WIN 35,065-2). Surprisingly, the 3alpha-phenyl isomers (6c, 6d) were more potent than the 3beta-phenyl isomers (6a, 6b). Molecular modeling studies revealed that the rigid 7-azabicyclo[2.2.1]heptane derivatives possess molecular topologies which are significantly different than the molecular topologies of the 2beta-(methoxycarbonyl)-3-phenyltropanes.

Structure-activity relationships at monoamine transporters and muscarinic receptors for N-substituted-3alpha-(3'-chloro-, 4'-chloro-, and 4',4''-dichloro-substituted-diphenyl)methoxytropanes.

The design, synthesis, and evaluation of 3alpha-(diphenylmethoxy)tropane (benztropine) analogues have provided potent and selective probes for the dopamine transporter. Structure-activity relationships (SARs) have been developed that contrast with those described for cocaine, despite significant structural similarity. Furthermore, behavioral evaluation of many of the benztropine analogues in animal models of cocaine abuse has suggested that these two classes of tropane-based dopamine uptake inhibitors have distinct pharmacological profiles. In general, the benztropine analogues do not demonstrate efficacious locomotor stimulation in mice, do not fully substitute for a cocaine discriminative stimulus, and are not appreciably self-administered in rhesus monkeys. These compounds are generally more potent than cocaine as dopamine uptake inhibitors in vitro, although their actions in vivo are not consistent with this action. These observations suggest that differing binding profiles at the serotonin and norepinephrine transporters as well as at muscarinic receptors might have significant impact on the pharmacological actions of these compounds. In addition, by varying the structures of the parent compounds and thereby modifying their physical properties, pharmacokinetics as well as pharmacodynamics will be directly affected. Therefore, in an attempt to systematically evaluate the impact of chemical modification on these actions, a series of N-substituted (H, CH3, allyl, benzyl, propylphenyl, and butylphenyl) analogues of 3'-chloro-, 4'-chloro-, and 4,4''-dichloro-3alpha-(diphenylmethoxy)tropanes were synthesized. These compounds were evaluated for displacement, in rat tissue, of [3H]WIN 35,428 from the dopamine transporter, [3H]citalopram from the serotonin transporter, [3H]nisoxetine from the norepinephrine transporter, and [3H]pirenzepine from muscarinic m1 receptors. SARs were developed and compared to a series of N-substituted-3alpha-(bis-4'-fluorophenyl)methoxytropanes. The present SARs followed previously reported studies with the single exception of the N-butylphenyl substituent, which did not provide the high affinity binding in any of these three sets of analogues, as it did in the 4',4''-difluoro series. X-ray crystallographic analyses of the three parent ligands (1a, 2a, and 3a) were compared to that of 3alpha-(bis-4'-fluorophenyl)methoxytropane which provided supportive evidence toward the proposal that the combination of steric bulk in both the 3-position and the N-substituent, in this class of compounds, is not optimal for binding at the dopamine transporter. These studies provide binding profile data that can now be used to correlate with future behavioral analyses of these compounds and may provide insight into the kind of binding profile that might be targeted as a potential treatment for cocaine abuse.