Profiling the expression of fatty acid-binding proteins and fatty acid transporters in mouse microglia and assessing their role in docosahexaenoic acid-d5 uptake.

While the processes governing docosahexaenoic acid (DHA) trafficking across the blood-brain barrier have been elucidated, factors governing DHA uptake into microglia, an essential step for this fatty acid to exert its anti-inflammatory effects, are unknown. This study assessed the mRNA and protein expression of fatty acid-binding proteins (FABPs) and fatty acid transport proteins (FATPs) in mouse BV-2 cells and their mRNA expression in primary mouse microglia. The microglial uptake of DHA-d5, a surrogate of DHA, was assessed by LC-MS/MS following interventions including temperature reduction, silencing of various FABP isoforms, competition with DHA, and metabolic inhibition. It was found that DHA-d5 uptake at 4°C was 39.6% lower than at 37°C, suggesting that microglial uptake of DHA-d5 likely involves passive and/or active uptake mechanisms. Of all FABP and FATP isoforms probed, only FABP3, FABP4, FABP5, FATP1, and FATP4 were expressed at both the mRNA and protein level. Silencing of FABP3, FABP4, and FABP5 resulted in no change in cellular DHA-d5 uptake, nor did concomitant DHA administration or the presence of 0.1% sodium azide/50 mM 2-deoxy-D-glucose. This study is the first to identify the presence of FABPs and FATPs in mouse microglia, albeit these proteins are not involved in the microglial uptake of DHA-d5.

Cue-conditioned alcohol seeking in rats following abstinence: involvement of metabotropic glutamate 5 receptors.

BACKGROUND AND PURPOSE: The current study was designed to: (i) examine whether functional interactions occur between receptors known to regulate alcohol self-administration; and (ii) characterize relapse to alcohol seeking following abstinence. EXPERIMENTAL APPROACH: The selective cannabinoid CB(1) receptor antagonist SR141716A (0.03-1.0 mg.kg(-1) i.p.) resulted in a dose-dependent reduction in ethanol self-administration in ethanol-preferring Indiana-preferring rats. SR141716A was then co-administered with either the selective glutamate metabotropic glutamate 5 (mGlu(5)) receptor antagonist 3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pyridine (MTEP) or the selective adenosine A(2A) receptor antagonist SCH58261. KEY RESULTS: When administered at individually sub-threshold doses, a combination of SR141716A (0.1 mg.kg(-1)) and SCH58261 (0.5 mg.kg(-1) i.p.) produced a reduction (28%) in ethanol self-administration. Combinations of threshold doses of SR141716A (0.3 mg.kg(-1)) and SCH58261 (2.0 mg.kg(-1), i.p.) caused an essentially additive reduction (68%) in alcohol self-administration. A combination of individually sub-threshold doses of CB(1) and mGlu(5) receptor antagonists did not affect alcohol self-administration; however, combined threshold doses of SR141716A (0.3 mg.kg(-1)) and MTEP (1.0 mg.kg(-1) i.p.) did reduce ethanol self-administration markedly (80%). Cue-conditioned alcohol seeking was attenuated by pretreatment with MTEP (1.0 mg.kg(-1)) co-administered with SR141716A (0.3 mg.kg(-1) i.p.). In contrast, SCH58261 (2.0 mg.kg(-1)) co-administered with SR141716A (0.3 mg.kg(-1) i.p.) did not reduce cue-conditioned alcohol seeking. CONCLUSIONS AND IMPLICATIONS: Adenosine A(2A) and cannabinoid CB(1) receptors regulated alcohol self-administration additively, but combined low-dose antagonism of these receptors did not prevent cue-conditioned alcohol seeking after abstinence. In contrast, combined low-dose antagonism of mGlu(5) and CB(1) receptors did prevent relapse-like alcohol seeking after abstinence, suggesting a prominent role for mGlu(5) receptors in this paradigm.

Genetic interdependence of adenosine and dopamine receptors: evidence from receptor knockout mice.

Dopamine and adenosine receptors are known to share a considerable overlap in their regional distribution, being especially rich in the basal ganglia. Dopamine and adenosine receptors have been demonstrated to exhibit a parallel distribution on certain neuronal populations, and even when not directly co-localized, relationships (both antagonistic and synergistic) have been described. This study was designed to investigate dopaminergic and purinergic systems in mice with ablations of individual dopamine or adenosine receptors. In situ hybridization histochemistry and autoradiography was used to examine the level of mRNA and protein expression of specific receptors and transporters in dopaminergic pathways. Expression of the mRNA encoding the dopamine D2 receptor was elevated in the caudate putamen of D1, D3 and A2A receptor knockout mice; this was mirrored by an increase in D2 receptor protein in D1 and D3 receptor knockout mice, but not in A2A knockout mice. Dopamine D1 receptor binding was decreased in the caudate putamen, nucleus accumbens, olfactory tubercle and ventral pallidum of D2 receptor knockout mice. In substantia nigra pars compacta, dopamine transporter mRNA expression was dramatically decreased in D3 receptor knockout mice, but elevated in A2A receptor knockout mice. All dopamine receptor knockout mice examined exhibited increased A2A receptor binding in the caudate putamen, nucleus accumbens and olfactory tubercle. These data are consistent with the existence of functional interactions between dopaminergic and purinergic systems in these reward and motor-related brain regions.

Characterisation of central adenosine A(1) receptors and adenosine transporters in mice lacking the adenosine A(2a) receptor.

The present study was designed to assess whether adenosine A(2a) receptor knockout mice exhibit altered purine utilisation in brain nuclei. Specifically, the properties of adenosine transporters and adenosine A(1) receptors were characterised in brain membranes and on slide-mounted sections. The B(MAX) for [(3)H]nitrobenzylthioinosine ([(3)H]NBTI) binding (adenosine transporter density) was significantly reduced in brainstem membranes of homozygotes (560+/-52 fmol/mg protein, n=5, P<0.05, Kruskal-Wallis ANOVA) compared to wildtype (1239+/-213 fmol/mg protein) and heterozygous mice (1300+/-558 fmol/mg protein). Quantitative autoradiography data indicated that [(3)H]NBTI binding in the medulla oblongata of heterozygous mice was seen to decrease significantly (P<0.05) in the subpostremal nucleus tractus solitarius (NTS), medial NTS, inferior olive and area postrema (AP). On the other hand, in the homozygous mice a decrease was seen in the medial NTS and AP. In the pons, [(3)H]1, 3-dipropyl-8-cyclopentylxanthine ([(3)H]DPCPX) (adenosine A(1) receptor density) binding increased significantly (P<0.05, Kruskal-Wallis ANOVA) in the lateral parabrachial nucleus, caudal pontine reticular nucleus and locus coeruleus of homozygotes compared to wildtype. In higher brain centres, [(3)H]NBTI binding was reduced in the paraventricular thalamic nucleus of both heterozygous and homozygous mice, whereas [(3)H]DPCPX binding was reduced in the hippocampus and lateral hypothalamus of heterozygotes. In homozygotes, [(3)H]DPCPX binding in the hippocampus increased compared to wildtype mice. The present study indicates that deletion of the A(2a) receptor may have contributed to region-specific compensatory changes in purine utilisation in brain nuclei associated with autonomic, neuroendocrine and behavioural regulation.

Visualisation of AMPA binding sites in the brain of mice lacking the adenosine A(2a) receptor.

Adenosine A(2a) receptor knockout mice (A(2a)R k/o) have been characterised as hypertensive, aggressive, anxious and hypoalgesic [12]. Thus, the present study was designed to investigate markers of glutamatergic transmission in specific brain nuclei associated with autonomic regulation. Visualisation of alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic acid binding sites in the brains of A(2a)R k/o mice was achieved by utilising quantitative receptor autoradiography with (S)-[(3)H]-5-fluorowillardiine ([(3)H]FW:10 nM) on slide-mounted sections of mouse-brain. [(3)H]FW binding significantly increased in the nucleus tractus solitarius and area postrema of A(2a)R k/o mice compared with wildtype CD-1 mice. In other autonomic nuclei examined, binding was unchanged between wildtype and knockout mice. The present study suggests that glutamatergic neurotransmission within certain neural pathways involved in autonomic and motor control is altered in the brains of A(2a)R k/o mice.

CCK/dopamine interactions in Fawn-Hooded and Wistar-Kyoto rat brain.

The aim of this study was to compare the actions of CCK neuropeptides within the nucleus accumbens (N.Acc) of alcohol preferring (Fawn-Hooded, FH) and alcohol nonpreferring (Wistar-Kyoto, WKY) rats. CCK-8S (30-300 nM) facilitated the K(+) stimulated release of [(3)H]dopamine (DA) from N.Acc prisms in both rat strains, whereas CCK-4 (30 nM-1 microM) caused a significant decrease of evoked [(3)H]DA in the FH rat only. A scattered distribution of CCK-A and -B receptor immunopositive varicose fibers were visualized throughout the N.Acc of both rat strains along with a topographic distribution of CCK receptor positive cells throughout the ventral mesencephalon.

Risk and protective factors for children's substance use and antisocial behavior following parental divorce.

Social networks and personal resources were examined as risk or protective factors for substance use and antisocial behavior in children five years after parental divorce. Children of divorce reported significantly more substance-using friends and less use of coping and social skills than children whose parents had not divorced. Findings suggest the importance of focusing on substance use as well as mental health outcomes in preventive interventions for children of divorce.

Evaluation of a preventive intervention for a self-selected subpopulation of children.

Evaluated an experimental preventive intervention developed for children who perceived their parents as problem drinkers. The 8-session program was designed to improve children's coping, self-esteem, and social competence, and modify alcohol expectancies which were specified as mediators of the effects of parental alcohol abuse on child mental health. Participants were 271 self-selected 4th-, 5th-, and 6th-grade students in 13 schools. The children were randomly assigned to treatment or delayed treatment conditions and the program was given to three successive cohorts of students. A meta-analysis across three different cohorts indicated significant program effects to improve knowledge of the program content and the use of support- and emotion-focused coping behaviors for the full sample. A slightly stronger range of effects was found for a high-risk subsample.

Development of a school-based prevention program for children in alcoholic families.

The systematic development of a preventive intervention for elementary-aged children of alcoholics (COAs) is described. First, the risk status of children of untreated alcoholics was established. Second, risk and protective factors that appeared to be mediators of mental health status for COAs were identified. Third, a preventive intervention was designed to teach coping skills and enhance self-esteem. Fourth, the intervention was pilot tested to assess its feasibility and potential. Finally, plans for a large scale experimental field trial of the revised curriculum are outlined. The advantages of following a systematic intervention development plan are demonstrated.

Vaginal wall sling.

We describe a new technique for the treatment of urinary incontinence due to intrinsic sphincteric damage in which a sling constructed from vaginal wall is used to provide compression and support of the urethra. A rectangular island of in situ anterior vaginal wall underlying the urethra and bladder neck is developed, the 4 corners are anchored with polypropylene sutures and a ligature carrier is used to transfer the sutures to a suprapubic location. An anterior vaginal wall flap proximal to the island is advanced to cover the island. When the sutures are tied the resulting sling will support the urethra and increase urethral resistance by compression, restoring continence. The advantages are its simplicity, need for only a small incision, short operative time and hospital stay, and reliance on healthy, well vascularized, in situ tissue. Continence has been achieved in 29 of 32 cases. All patients voided spontaneously except for those with neuropathic urethral incompetence who required self-catheterization.

Risk status of adolescent children of problem-drinking parents.

These studies assessed the risk status of children of untreated alcoholics. In Study 1, a cross-sectional survey of 208 high school students identified 18% as having serious concern about their parents' drinking. In Study 2, 32 children of problem-drinking parents and 39 others who participated in self-help groups were surveyed. Children of problem-drinking parents were more at risk of depression, low self-esteem, and heavy drinking than their peers in the general high school population. Within self-help groups, however, children's symptomatology was not related to their parents' drinking status. The results support the need for preventive intervention for children of problem-drinking parents and for developing strategies for improving the participation rate of such children in the intervention programs that are available.

Reduction of Lactic Acid, Nonprotein Nitrogen, and Ash in Lactic Acid Whey by Candida ingens Culture.

A simple, efficient procedure for removing lactic acid and for reducing nonprotein nitrogen and ash in lactic acid whey has been developed. The procedure consists of culturing Candida ingens on the whey. This organism could assimilate >98% of the lactic acid and approximately 40% of the nonprotein nitrogen. Ash reduction of up to 45% resulted from precipitation of calcium apatite due to the increase in pH from 4.4 to approximately 8.0 which occurred during growth of C. ingens. Improved fluxes during laboratory-scale ultrafiltration were obtained for the treated lactic acid whey. C. ingens treatment of lactic acid whey appears to facilitate processing of this material to a more useful product.