Harm reduction has gradually entered social work discourse and is now seen as a promising approach for treating individuals with drug and alcohol problems. However, beyond statements and data ...supporting the utility of a harm reduction approach, few guidelines for clinical practice have been detailed in the social work literature. This lack of concrete detail regarding how harm reduction is actually practiced limits the potential implementation of the model into day-to-day clinical work. This article reiterates that harm reduction is a viable approach to clinical social work practice with individuals who have drug- and alcohol-related problems and for whom traditional approaches may be inappropriate. It focuses on harm reduction therapy as an emerging treatment model that can be implemented by clinical social workers and mental-health and substance use treatment providers. The article identifies and elaborates several basic tenets that can be incorporated into clinical social work. It is hoped that social workers who learn how harm reduction is implemented in clinical practice will be more apt to incorporate its principles into their work.
The binding site for DETQ 2-(2,6-dichlorophenyl)-1-((1
,3
)-3-(hydroxymethyl)-5-(2-hydroxypropan-2-yl)-1-methyl-3,4-dihydroisoquinolin-2(1
)-yl)ethan-1-one, a positive allosteric modulator (PAM) of ...the dopamine D1 receptor, was identified and compared with the binding site for CID 2886111
-(6-
-butyl-3-carbamoyl-4,5,6,7-tetrahydro-1-benzothiophen-2-yl)pyridine-4-carboxamide, a reference D1 PAM. From D1/D5 chimeras, the site responsible for potentiation by DETQ of the increase in cAMP in response to dopamine was narrowed down to the N-terminal intracellular quadrant of the receptor; arginine-130 in intracellular loop 2 (IC2) was then identified as a critical amino acid based on a human/rat species difference. Confirming the importance of IC2, a
2-adrenergic receptor construct in which the IC2 region was replaced with its D1 counterpart gained the ability to respond to DETQ. A homology model was built from the agonist-state
2-receptor structure, and DETQ was found to dock to a cleft created by IC2 and adjacent portions of transmembrane helices 3 and 4 (TM3 and TM4). When residues modeled as pointing into the cleft were mutated to alanine, large reductions in the potency of DETQ were found for Val119 and Trp123 (flanking the conserved DRY sequence in TM3), Arg130 (located in IC2), and Leu143 (TM4). The D1/D5 difference was found to reside in Ala139; changing this residue to methionine as in the D5 receptor reduced the potency of DETQ by approximately 1000-fold. None of these mutations affected the activity of CID 2886111, indicating that it binds to a different allosteric site. When combined, DETQ and CID 2886111 elicited a supra-additive response in the absence of dopamine, implying that both PAMs can bind to the D1 receptor simultaneously.
The hemagglutinin-tagged human trace amine-associated receptor1 (TAAR1) was stably coexpressed with rat Galpha(s) in the AV12-664 cell line, and receptor activation was measured as the stimulation of ...cAMP formation. After blockade of endogenously expressed alpha2- and beta-adrenoceptors with 2-2-(2-methoxy-1,4-benzodioxanyl)-imidazoline hydrochloride (2-methoxyidazoxan, RX821002) and alprenolol, respectively, the resulting pharmacology was consistent with that of a unique receptor subtype. beta-Phenylethylamine (beta-PEA), the putative endogenous ligand, gave an EC50 of 106 +/- 5 nM in the assay. For a series of beta-PEA analogs used to explore the pharmacophore, small substituents at ring positions 3 and/or 4 generally resulted in compounds having lower potency than beta-PEA, although several were as potent as beta-PEA. However, small substituents at ring position 2 resulted in a number of compounds having potencies as good as or better than beta-PEA. A number of nonselective antagonists known to share affinity for multiple monoaminergic receptors were evaluated for their ability to inhibit beta-PEA stimulation of the human TAAR1. None had an IC50 <10 microM. For comparison, the rat TAAR1 receptor was expressed in the AV12-664 cell line. A number of agonist compounds had significantly different relative potencies between the rat and human TAAR1, demonstrating a significant species difference between the rat and human TAAR1. The TAAR1 receptor exhibits a pharmacologic profile uniquely different from those of classic monoaminergic receptors, consistent with the structural information that places them in a distinct family of receptors. This unique pharmacologic profile suggests the potential for development of TAAR-selective agonists and antagonists to study their physiologic roles.
Human kallikrein 6 (hK6) is a trypsin-like serine protease, member of the human kallikrein gene family. Studies suggested a potential involvement of hK6 in the development and progression of ...Alzheimer’s disease. The serum levels of hK6 might be used as a biomarker for ovarian cancer. To gain insights into the physiological role of this enzyme, we sought to determine its substrate specificity and its interactions with various inhibitors. We produced the proform of hK6 and showed that this enzyme was able to autoactivate, as well as proteolyse itself, leading to inactivation. Kinetic studies indicated that hK6 cleaved with much higher efficiency after Arg than Lys and with a preference for Ser or Pro in the P2 position. The efficient degradation of fibrinogen and collagen types I and IV by hK6 indicated that this kallikrein might play a role in tissue remodeling and/or tumor invasion and metastasis. We also demonstrated proteolysis of amyloid precursor protein by hK6 and determined the cleavage sites at the N-terminal end of the protein. Inhibition of hK6 was achieved via binding to different serpins, among which antithrombin III was the most efficient.
Human kallikrein 7 (hK7), also known as human stratum corneum chymotryptic enzyme, is a chymotrypsin-like serine protease first identified in human skin extracts and predicted to be a secreted ...protease. The aim of this study was to develop a sensitive and specific immunoassay for hK7 and to examine the distribution of hK7 in tissue extracts and biological fluids.
Recombinant hK7 was produced in human embryonic kidney cells (HEK293T) and purified by a three-step column chromatographic procedure. The purified hK7 was injected into mice for antibody generation. A sandwich-type immunoassay was developed with the anti-hK7 monoclonal antibodies.
The assay had imprecision (CV) <10% through the dynamic range of 0.2-20 microg/L and had no detectable cross-reactivity from other members in the human kallikrein gene family. Highest concentrations were found in skin, esophagus, and kidney. hK7 was also found in amniotic fluid, ascites from ovarian cancer patients, breast milk, cerebrospinal fluid, saliva, seminal plasma, serum, sweat, synovial fluid, and urine.
This study describes the first ELISA-type immunoassay for hK7 protein quantification. hK7 is found many human tissues and in various biological fluids.
The human 5-HT(1E) receptor gene was cloned more than a decade ago. Little is known about its function, and there have been no reports of its existence in the genome of small laboratory animals. In ...this study, attempts to clone the 5-HT(1E) gene from the rat and mouse were unsuccessful. In fact, a search of the mouse genome database revealed that the 5-HT(1E) receptor gene is missing from the mouse genome. However, the 5-HT(1E) gene was cloned from guinea pig genomic DNA and was characterized. The guinea pig 5-HT(1E) receptor gene encodes a protein of 365 amino acids. It shares 88% (nucleic acid) and 95% (amino acid) homology with the human receptor. The guinea pig 5-HT(1E) receptor showed similar pharmacology to the human 5-HT(1E) receptor in radioligand binding assays. Serotonin (5-hydroxytryptamine, 5-HT) dose-dependently stimulated 35SGTPgammaS binding to the guinea pig 5-HT(1E) receptor with an EC(50) of 13.6+/-1.92 nM, similar to that of the human 5-HT(1E) receptor (13.7+/-1.78 nM). Activation of the guinea pig 5-HT(1E) receptor was also achieved by ergonovine, alpha-methyl-5-HT, 1-naphthylpiperazine, methysergide, tryptamine, and 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI). Methiothepin exhibited antagonist activity. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that 5-HT(1E) mRNA was present in the guinea pig brain with the greatest abundance in the hippocampus, followed by the olfactory bulb. Lower levels were detected in the cortex, thalamus, pons, hypothalamus, midbrain, striatum, and cerebellum. Our current study marks the first identification of the 5-HT(1E) receptor gene in a commonly used laboratory animal species. This finding should allow the elucidation of the receptor's role(s) in the complex coordination of central serotonergic effects.
The KLK6 gene is a new member of the human kallikrein gene family and encodes for a secreted protease, human kallikrein 6 (hK6; also known as zyme/protease M/neurosin). No study has as yet reported ...detailed immunohistochemical localization of hK6 in human tissues. Our purpose was to examine the expression of hK6 in human tissues by immunohistochemistry. We have analyzed 199 paraffin blocks from archival, current, and autopsy material prepared from almost every normal human tissue. We employed an hK6-specific polyclonal rabbit antibody and avidin-biotin to localize hK6 by IHC. The staining pattern, the distribution of the immunostaining, and its intensity were studied in detail. The IHC expression of zyme was generally cytoplasmic. Various normal human tissues expressed the protein abundantly. Glandular epithelia constituted the main immunoexpression sites, with representative organs being the breast, prostate, kidney, endometrium, colon, appendix, salivary glands, bile ducts, and gallbladder. The small intestine, stomach, endocervix, Fallopian tube, epididymis, bronchus, and upper respiratory tract showed a focal expression as well. Choroid plexus epithelium, peripheral nerves, and some neuroendocrine cells (including the islets of Langerhans, cells in the anterior pituitary gland, and adrenal medulla) expressed the protein strongly and diffusely. A characteristic immunostaining was observed in the Hassall's corpuscles of the thymus, the oxyphilic cells of the thyroid and parathyroid glands, the primordial follicles of the ovary, dendritic cells mainly in the spleen, and in various cells of the placenta.
The dopamine D1 receptor agonist dihydrexidine (DHX) (±)-trans-10,11-dihydroxy-5,6,6a,7,8,12b-hexahydrobenzoa phenanthridine hydrochloride has shown efficacy in animal models of Parkinsonʼs disease ...and improved cerebral blood flow and working memory of schizophrenic patients. Although the discriminative stimulus effects of DHX, an in-vivo predictor of human subjective effect profile, have only been characterized with respect to activity at D1 receptors, DHX also has significant affinity for D2 receptors. This study was designed to characterize the role of D1 and D2/D3 receptors in mediating the discriminative stimulus effects of DHX. Rats were trained to discriminate DHX 3 mg/kg, intraperitoneally (i.p.) from the vehicle. The selective dopamine D1 receptor partial agonist SKF 38393 was fully substituted for DHX. The D1 receptor antagonist SCH 23390 (0.1 mg/kg, s.c.) and the D3-selective antagonist U99194 (10 mg/kg, i.p.) significantly attenuated the discriminative stimulus effects of the training dose of DHX by 80 and 60%, respectively, suggesting that both D1 and D3 receptors mediate the discriminative stimulus effects of DHX. In contrast, raclopride (1 mg/kg, i.p.) did not significantly alter the discriminative stimulus effects of DHX, indicating a lack of D2-mediated effects. The D2/D3 receptor preferring agonists, quinpirole and (+)-PD 128907 were fully substituted, whereas (+)-7-OH-DPAT was partially substituted for DHX. The DHX bound to D2 receptors with a Ki of 4.3+0.7 nmol/l was compared with 33.7+4.6 nmol/l at D3 receptors. Determinations of activity at second messenger systems revealed that DHX functioned as a full agonist at D3 receptors and a partial agonist at D2 receptors in vitro. These activities at D2/D3 receptors have shown effects in some preclinical models and clinical disease states. Therefore, the prominent in-vivo agonist activity of DHX at both D1 receptors and D2/D3 receptors should be considered while making predictions of effects in humans.
Phospholipid breakdown has been reported to be an early event in the brain after global cerebral ischemia. Our earlier observations showing the localization of cytosolic phospholipase A2 (cPLA2) to ...astrocytes in aged human brains and the intense glial activation observed after global forebrain ischemia prompted us to investigate the cellular localization of cPLA2 in the rat brain subjected to global ischemia.
Immunohistochemistry was performed in sections through the dorsal hippocampus in rats subjected to 30 minutes of four- vessel occlusion. PLA2 was localized with the use of a highly selective antiserum. Double immunofluorescent localization was performed to colocalize cPLA2 with various glial cell types. cPLA2 levels were also measured by enzymatic assay and Western blot analysis.
A marked induction of cPLA2 was observed in activated microglia and astrocytes in the CA1 hippocampal region at 72 hours after ischemia. Only a subset of astrocytes and microglia were immunoreactive for cPLA2. Twenty-four hours after ischemia, numerous cPLA2 immunoreactive astrocytes were observed. Western blot analysis of hippocampal homogenates at 72 hours after ischemia showed induction of a 100-kD band that comigrated with purified human cPLA2, and a threefold induction in cPLA2 activity was demonstrated by enzymatic assay.
These results indicate that both reactive astrocytes and microglia contain elevated levels of cPLA2. Induction of cPLA2 was confined to areas of neurodegeneration and likely precedes its onset. The results suggest that reactive glia may play a role in the pathophysiology of delayed neuronal death after transient global forebrain ischemia.
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