Advances in pharmacology (San Diego, Calif.)

Published by Elsevier
Online ISSN: 1054-3589
Publications
Plasma concentration profiles of pramlintide following intravenous infusion of doses of 30, 100, and 300 mg over 2 min (left panel) or 2 hr (right panel) Data from Colburn et al. (1996).
Reduction in 24-hr glucose profiles with 30 mg pramlintide administered four times daily to subjects with type 1 diabetes. Data from Thompson et al. (1997).
Effect of pramlintide on plasma fructosamine in patients with type 1 (Thompson et al., 1997) and type 2 diabetes (Thompson et al., 1998).
Effect of pramlintide on body weight in patients with type 1 diabetes. Weight loss was greatest in those with higher BMI. Data from Ratner et al. (2004).
Effects of pramlintide infused at 100 mg/hr on glucose and glucagon profiles following a test meal in patients with type 2 diabetes treated with combined insulin þ sulfonylurea (upper panel), insulin alone (middle panel), or sulfonylurea alone (lower panel). Data from Fineman et al. (2002).
Article
Recognizing type 1 diabetes is characterized not only by insulin deficiency, but also by amylin deficiency. Cooper predicted that certain features of the disease could be related and he proposed the amylin/insulin coreplacement therapy. Although the early physiological rationale was flawed, the idea that glucose control could be improved over that attainable with insulin alone without invoking the ravages of worsening insulin-induced hypoglycemia was vindicated. The principal risk of pramlintide cotherapy is an increased probability of insulin-induced hypoglycemia, especially at the initiation of therapy. This risk could be mitigated by preemptive reduction in insulin dose. Effects observed in animals, such as the slowing of gastric emptying, the inhibition of nutrient-stimulated glucagon secretion, and the inhibition of food intake, generally have been replicated in humans. A notable exception appears to be the induction of muscle glycogenolysis and an increase in plasma lactate.
 
Article
This chapter presents evidence that calpain over activation may be a key component in a number of disorders. The common theme for most of these disorders is cellular Ca2+ overload. The calpains can be found in the cytosolic compartment and can also be identified in the plasma membrane and other organelle membranes. Both isoforms contain two subunits. Calpain appears to be an ideal pharmaceutical target, as this protease is most active during pathological events. The current challenge is to identify cell-permeable and selective calpain inhibitors for evaluation in various in vivo disease models. As the knowledge of calpain substrate specificity is increasing and calpain inhibitors from natural products screening programs are becoming available, more selective inhibitors are designed and synthesized. At present there is an arsenal of calpain inhibitors for disposal. These inhibitors include protein inhibitors, such as calpastatin, irreversible peptide inhibitors, such as E64 analogs, reversible peptide inhibitors, such as peptidyl α-keto amides, and several nonpeptide inhibitors. With the recent emergence of several cell-permeable calpain selective inhibitors, the goal of understanding the physiological roles of calpain are possible to be achievable.
 
Article
Studies have shown that the dopaminergic neurons terminating in the intermediate lobe of the pituitary (ILP) originate in the periventricular nucleus of the hypothalamus. A recent observation revealed that tyrosine hydroxylase (TH) colocalized with calretinin (CR), a calcium- binding protein, in the rat ILP. In addition, TH also colocalized with calbindin-D28k (CB), another calcium-binding protein (CaBP). The purpose of this chapter is to determine, using an immunohistochernical colocalization procedure, whether a triple colocalization of TH, CR, and CB existed in the DA fibers of the ILP. As it is already demonstrated that such a triple colocalization exists, the chapter sets out to map the dopaminergic cells in the rat hypothalamus in a search for the origin of these pituitary fibers. In immunocytochemical preparations of rat pituitary glands, varicose dopaminergic nerve fibers formed a plexus enveloping the endocrine cells of the intermediate lobe, and the lack of anti-DBH staining confirmed the DA nature of these fibers. Triple colocalization of TH, CR, and CB revealed that all three neurochemicals were present in these fibers in what appeared to be a 1:1 relationship between the CaBPs (CR, CB) and the TH-containing fibers. No immunoreactive cell bodies were observed in the ILP. Anatomical, neurochemical, and neuroendocrinological evidence indicates that dopamine terminals in the ILP originate from A14 cells in the hypothalamic periventricular nucleus. It has been observed that a triple colocalization of TH, CR, and CB was contained in about 3.3% of the A14 DA cell bodies, and it is suggested that these are the cells that innervate the ILP. Because of the triple colocalization within approximately 3.1% of the A11 cells, this system may also project to the ILP.
 
Article
The P2Y receptors are a widely expressed group of eight nucleotide-activated G protein-coupled receptors (GPCRs). The P2Y(1)(ADP), P2Y(2)(ATP/UTP), P2Y(4)(UTP), P2Y(6)(UDP), and P2Y(11)(ATP) receptors activate G(q) and therefore robustly promote inositol lipid signaling responses. The P2Y(12)(ADP), P2Y(13)(ADP), and P2Y(14)(UDP/UDP-glucose) receptors activate G(i) leading to inhibition of adenylyl cyclase and to Gβγ-mediated activation of a range of effector proteins including phosphoinositide 3-kinase-γ, inward rectifying K(+) (GIRK) channels, phospholipase C-β2 and -β3, and G protein-receptor kinases 2 and 3. A broad range of physiological responses occur downstream of activation of these receptors ranging from Cl(-) secretion by epithelia to aggregation of platelets to neurotransmission. Useful structural models of the P2Y receptors have evolved from extensive genetic analyses coupled with molecular modeling based on three-dimensional structures obtained for rhodopsin and several other GPCRs. Selective ligands have been synthesized for most of the P2Y receptors with the most prominent successes attained with highly selective agonist and antagonist molecules for the ADP-activated P2Y(1) and P2Y(12) receptors. The widely prescribed drug, clopidogrel, which results in irreversible blockade of the platelet P2Y(12) receptor, is the most important therapeutic agent that targets a P2Y receptor.
 
Article
The authors have presented a concise review of the studies which evaluate the risk of colorectal cancer among NSAID users. Animals studies have clearly documented a protective effect of NSAIDs in preventing colon cancers in a carcinogen-induced (AOM) model. NSAIDs are protective in the animal model, even if given 14 weeks after administration of the carcinogen, indicating that they must be playing a role very early in the adenoma-to-carcinoma sequence of events. Several studies have indicated that treatment of FAP patients with NSAIDs causes a regression of adenomas that were already present prior to initiation of NSAID therapy. Many epidemiological studies have examined the relationship between aspirin use and colorectal cancer. Most of these studies have shown a marked decrease in the relative risk (40-50%) of colorectal cancer among continuous aspirin users. The appropriate dose and duration of aspirin treatment for optimal effects are still unknown. Future work, directed at the molecular basis for the chemoprotective effects of NSAIDs in humans, may reveal strategies for the development of better chemopreventive agents. One effect shared by all NSAIDs is their ability to inhibit cyclooxygenase. Presently, it is not clear whether inhibition of cyclooxygenase-1 or -2 effects on other signaling pathways are required for the protective effect of aspirin and other NSAIDs. The authors and others have demonstrated that COX-2 is upregulated from 2- to 50-fold in 85-90% of colorectal adenocarcinomas, which makes the COX-2 enzyme a possible target. Drugs are currently under development at several pharmaceutical companies that preferentially inhibit either COX-2 or COX-2. If COX-2 is found to be a relevant target in the prevention of colorectal cancer, then these newly developed, more selective NSAIDs may play a role in future chemoprevention strategies.
 
Article
Islet cell autoantigen (ICA) 512 is a specific membrane marker of neurosecretory granules. It is the first member of the receptor protein tyrosine phosphatases (RPTPs) family found to be resident in an intracellular compartment. Interestingly, ICA 512 contains only one protein tyrosine phosphatase (PTP) “core domain”, rather than two, as usually found in receptor PTPs (RPTPs). Moreover, ICA 512 expressed in bacteria do not display PTP activity when tested with several common PTP substrates. Thus, the function of ICA 512 is still unknown. By Northern blot, ICA 512 transcripts have been detected in pancreatic islets, brain, and pituitary, suggesting an enrichment of ICA 512 in neuroendocrine tissues. To begin addressing the structure and function of ICA 512, its tissue distribution and intracellular localization using rabbit antibodies directed against either its recombinant extracellular domain or recombinant cytoplasmic domain. Confocal microscopy on rat tissues demonstrated that ICA 512 is expressed in virtually all neuroendocrine cells, including neurons of the autonomic nervous system; chromaffin cells of the adrenal medulla; α-, β-, and δ-pancreatic islet cells; cells in the anterior and intermediate pituitary, as well as neurons of the autonomic nervous system and of the hypothalamus and the amygdala in the brain. The strongest ICA 512 immunoreactivity has been found in the posterior pituitary, which contains the nerve endings of neurons located in the hypothalamus and the highest concentration of neurosecretory granules in the entire body. Thus ICA 512 and, thus, tyrosine phosphorylation–dephosphorylation plays a role in the life-cycle of neurosecretory granules.
 
Chapter
According to The European Monitoring Centre for Drugs and Drug Addiction (EMCDDA), ∼70 million European adults have consumed cannabis on at least one occasion. Cannabis consumption leads to a variety of psychoactive effects due to the presence of the constituent Δ(9)-tetrahydrocannabinol (Δ(9)-THC). Δ(9)-THC interacts with the endocannabinoid system (ECS), which consists of the seven transmembrane spanning (7TM)/G protein-coupled receptors (GPCRs) CB(1) and CB(2), their respective ligands (endocannabinoids), and enzymes involved in their biosynthesis and degradation. This system plays a critical role in many physiological processes such as learning and memory, appetite control, pain sensation, motor coordination, lipogenesis, modulation of immune response, and the regulation of bone mass. Therefore, a huge effort has been spent trying to fully elucidate the composition and function of the ECS. The G protein-coupled receptor 55 (GPR55) was recently proposed as a novel component of this system; however, its classification as a cannabinoid receptor has been significantly hampered by its complex pharmacology, signaling, and cellular function. GPR55 is phylogenetically distinct from the traditional cannabinoid receptors, but in some experimental paradigms, it is activated by endocannabinoids, phytocannabinoids, and synthetic cannabinoid ligands. However, the most potent compound appears to be a lysophospholipid known as lysophosphatidylinositol (LPI). Here, we provide a comprehensive evaluation of the current pharmacology and signaling of GPR55 and review the proposed role of this receptor in a number of physiological and pathophysiological processes.
 
Article
It is known that Ca2+ is an important intracellular second messenger. Ca2+ binds to a variety of proteins and alters their functions. Calmodulin (CaM) is a ubiquitous Ca2+-binding protein and the Ca2+/CaM complex interacts with target proteins, resulting in their functional changes. In this respect, it is believed that the family of Ca2+/CaM-dependent protein kinases (CaM kinases) is important for Ca2+-mediated protein phosphorylation. The prototype of this family is smooth muscle myosin light-chain kinase (MLCK), which phosphorylates myosin regulatory light chain in a Ca2+/CaM-dependent manner resulting in the activation of actomyosin ATPase required for muscle fiber contraction. The multifunctional CaM kinases have different tissue distributions and substrate specificities, suggesting that each of the enzymes is involved in different types of Ca2+-mediated processes. Researchers have succeeded in synthesizing the potent, specific inhibitor of CaM kinase II, KN-62 (1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-l-tyrosyl)-4-phenylpiperazine). CaM kinase II is distributed in a wide range of tissues, but the brain contains the highest amount. This chapter reviews previous studies with KN-62 and propose the involvement of CaM kinase II in various aspects of cell function. Researchers have succeeded in synthesizing W-7, a naphthalenesulfonamide derivative, which can inhibit a protein activator-stimulated cyclic nucleotide phosphodiesterase (PDE). When the naphthalene ring was replaced by an isoquinoline ring in the derivatives with a shorter alkyl chain, the isoquinolinesulfonamide derivatives were no longer CaM antagonists but potent protein kinase inhibitors. KN-62 was proposed to be a specific inhibitor of CaM kinase II because the compound could selectively inhibit the enzyme among a limited number of protein kinases such as PKA, PKC, and smooth muscle MLCK. Research findings suggest that KN-62 occupies the Ca2+/CaM binding domain of CaM kinase II, thereby inhibits its Ca2+/CaM-dependent activation.
 
Article
Drugs that enhance the release of neurotransmitters, or that otherwise affect neurotransmission via a presynaptic action have potential use in a wide range of neurological diseases, including perhaps Alzheimer's disease, Parkinsonism, and depression. The discovery of linopirdine [DuP 996]; 3,3-bis(4-pyridinylmethyl)-1-phenylindolin-2-one; AVIVATM offered a new approach to the improvement of cholinergic function, and the drug has subsequently been used in clinical trials in Alzheimer's patients. Linopirdine was found to increase the release of ACh and other neurotransmitters in response to a depolarizing stimulus. By facilitating neurotransmission only during neuronal firing, it might be expected that the functioning of impaired cholinergic pathways in the CNS could be improved. There is no reason to suppose that the cholinergic approach to the treatment of Alzheimer's disease would reverse the gross histological manifestations of the disease: the neurofibrillary tangles and neuritic (senile) plaques. Continued studies on the etiology of Alzheimer's are clearly of the utmost importance in determining new strategies for drug treatment, and possibly also for prevention. The nature of the disease causes it to severely impact the families of patients, as well as the patients themselves, and it is currently the fourth leading cause of death in the United States. Clinical management of Alzheimer's disease is handicapped by the heterogeneity of the illness, and by the difficulty of detecting it in the earlier stages. This chapter reviews the pharmacological actions of linopirdine with a particular emphasis on understanding the mechanism by which these effects are achieved. It is hoped that a better understanding of linopirdine's action can lead to new improved compounds, and perhaps can suggest new target sites for the treatment of Alzheimer's disease and other neurological disorders.
 
Article
A substantial amount of research has been applied to Phospholipase A2 (PLA2) enzymes, from a mechanistic understanding of enzyme structure function to the design of specific inhibitors for the treatment of inflammatory disorders because PLA2-catalyzed release of arachidonic acid (AA) is believed to be the rate-limiting event in the generation of proinflammatory lipid mediators. This chapter discusses selected classes of PLA2 inhibitors because these have generated the most intense research in the field. Investigation revealed that there is a multitude of structurally diverse compounds that are inhibitors of PLA2 in vitro and some of these compounds have antiinflammatory activity. This may be because of the nature of these inhibitors, as they are hydrophobic they may be more readily absorbed in the skin whereas when given orally they may not be absorbed. The concept of regulation of eicosanoid biosynthesis by PLA2 inhibition and decreased AA availability still remains a viable therapeutic approach for the treatment of inflammatory diseases. The proof of this concept has not been obtained because of the complex nature of PLA2 and the multiple forms of PLA2 in the cell. Clinical results with cyclooxygenase inhibitors and recent clinical results with inhibitors of 5-lipoxygenase prove that if inhibition of PLA2 results in reduction in both lipid mediators, a good anti-inflammatory compound results. The added advantage of PLA2 inhibitors would be the reduction in PAF levels; however, the clinical results with potent and specific PAF antagonists has been less encouraging about the potential benefits of reduction in PAF levels. Focused research, on the regulation of AA release from cells and the roles of different isoforms of PLA2 in the regulation of AA release have the potential to generate good anti-inflammatory compounds.
 
Article
Recent progress in our understanding of the unique role of A(2B) receptors in the regulation of inflammation, immunity, and tissue repair was considerably facilitated with the introduction of new pharmacological and genetic tools. However, it also led to seemingly conflicting conclusions on the role of A(2B) adenosine receptors in inflammation with some publications indicating proinflammatory effects and others suggesting the opposite. This chapter reviews the functions of A(2B) receptors in various cell types related to inflammation and integrated effects of A(2B) receptor modulation in several animal models of inflammation. It is argued that translation of current findings into novel therapies would require a better understanding of A(2B) receptor functions in diverse types of inflammatory responses in various tissues and at different points of their progression.
 
Article
That adenosine signaling can elicit adaptive tissue responses during conditions of limited oxygen availability (hypoxia) is a long-suspected notion that recently gained general acceptance from genetic and pharmacologic studies of the adenosine signaling pathway. As hypoxia and inflammation share an interdependent relationship, these studies have demonstrated that adenosine signaling events can be targeted to dampen hypoxia-induced inflammation. Here, we build on the hypothesis that particularly the A(2B) adenosine receptor (ADORA(2B)) plays a central role in tissue adaptation to hypoxia. In fact, the ADORA(2B) requires higher adenosine concentrations than any of the other adenosine receptors. However, during conditions of hypoxia or ischemia, the hypoxia-elicited rise in extracellular adenosine is sufficient to activate the ADORA(2B). Moreover, several studies have demonstrated very robust induction of the ADORA(2B) elicited by transcriptional mechanisms involving hypoxia-dependent signaling pathways and the transcription factor "hypoxia-induced factor" 1. In the present chapter, genetic and pharmacologic evidence is presented to support our hypothesis of a tissue protective role of ADORA(2B) signaling during hypoxic conditions, including hypoxia-elicited vascular leakage, organ ischemia, or acute lung injury. All these disease models are characterized by hypoxia-elicited tissue inflammation. As such, the ADORA(2B) has emerged as a therapeutic target for dampening hypoxia-induced inflammation and tissue adaptation to limited oxygen availability.
 
Article
Aromatic l-amino acid decarboxylase (AADC) is a homodimeric pyridoxal phosphate-dependent enzyme responsible for the syntheses of dopamine and serotonin. Defects in the AADC gene result in neurotransmitter deficiencies. Patients with AADC deficiency have severe motor and autonomic dysfunctions. A mouse model of AADC deficiency was recently established. These mice grow poorly and move awkwardly during infancy. They also show high anxiety when they grow up. Because drug therapy provides little or no benefit for many patients with AADC deficiency, a gene therapy has been attempted. The gene therapy employed an adeno-associated virus viral vector that can express the human AADC protein. The vector was injected to the brain of several children with AADC deficiency. The therapy was well tolerated, and all treated patients showed improvement. In the future, the mouse model will also help the development of treatments for AADC deficiency.
 
Article
It is now known that most of the drug-metabolizing enzyme (DME) systems present in the human liver, for example, cytochrome P450, CYP; UDP-glucuronosyltransferase, UDPGT; Nacetyltransferase, NAT; and NADPH-dependent flavin-containing monooxygenase, FMO are composed of two or more enzymes or “isforms” that can differentially interact with any number of drugs. A well-managed drug-drug interaction, as in the case of ketoconazole and cyclosporin, can potentially be of economic benefit to the patient. Likewise, a potent interaction with an enzyme such as CYP can lead to autoinhibition of metabolism and superior pharmacokinetics. In vitro drug-drug interaction screening is fast becoming a standardized part of the drug development process and, depending on the particular pharmaceutical company and/or NDE, can be performed prospectively, concurrently, or respectively. In most cases, in vitro studies focus on the human liver microsomal CYP “superfamily” of proteins, as our present understanding of this system exceeds that of other DME systems. As the procedures for phenotyping and/or genotyping of clinical subjects become more readily available, in vivo confirmation of the polymorphic oxidations will be greatly facilitated. There is no all-encompassing in vitro and/or in vivo animal model for predicting drug-drug interactions in humans. Because the liver is considered to be the major site of metabolism, most investigators now find themselves routinely performing in vitro human metabolism studies with various combinations of primary cultured hepatocytes, precision-cut liver slices, banks of liver microsomes, and/or cDNA-expressed enzymes.
 
Article
Brain capillary endothelial cells express multiple ATP-binding cassette transport proteins on the luminal, blood-facing, plasma membrane. There these transporters function as ATP-driven efflux pumps for xenobiotics and endogenous metabolites, providing an important element of the barrier. When the transporters limit neurotoxicant entry into the central nervous system (CNS), they are neuroprotective; when they limit therapeutic drug entry, they are obstacles to drug delivery to treat CNS diseases. Certainly, changes in the transporter expression and transport activity can have a profound effect on CNS pharmacotherapy, with increased transport activity reducing drug access to the brain and vice versa. Here, I review the signals that alter transporter expression and transport function with an emphasis on P-glycoprotein, MRP2, and breast cancer resistance protein (ABCG2) (BCRP), the efflux transporters for which we have the most detailed picture of regulation. Recent work shows that transporter protein expression can be upregulated in response to inflammatory and oxidative stress, therapeutic drugs, diet, and persistent environmental pollutants; as a consequence, drug delivery to the brain is reduced. For many of these stimuli, the transcription factor, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), appears to be involved. However, NF-κB activation and nuclear translocation is often initiated by upstream signaling. In contrast, basal transport activity of P-glycoprotein and BCRP can be reduced through complex signaling pathways. Targeting such signals provides opportunities to rapidly and reversibly increase brain accumulation of drugs that are transporter substrates. The extent to which such signaling-based strategies can be utilized in the clinic remains to be seen.
 
Article
Apoptosis is a genetically controlled process and is probably the counterbalancing force to mitosis under normal physiological conditions. About apoptosis it is known that the cell plays an active part in its own demise. A variety of genes that play regulatory roles in apoptosis have been identified. These fall into two distinct groups—those that drive the process and those that inhibit it. In the latter category are genes such as bcl-2 and bcr-abl. Many of these genes are oncogenes and have been implicated in the development of numerous cancers. This chapter focuses on a particular gene and how it contributes to the development of a particular form of cancer by interfering with the normal regulation of apoptosis. The chapter also describes the current understanding of the role played by Abl tyrosine kinases in regulating apoptosis, with particular reference to chronic myelogenous leukemia (CML). CML is a hematopoietic disorder that is characterized by the presence of excessive numbers of mature myeloid cells in the peripheral blood. Cancers in general are associated with consistent chromosomal abnormalities, and CML is the first human cancer to be associated with such an abnormality. The cytological hallmark of CML is the presence of the Philadelphia (Ph) chromosome, which results from a reciprocal translocation of c-abl from chromosome 9 to the breakpoint cluster region (bcr) gene on chromosome 22, generating the bcr-abl gene. A better understanding of the basic biology of Abl should enable the design of more specific inhibitors of the enzyme and the design of more effective therapeutic strategies for the long-term treatment of CML.
 
Article
The locus coeruleus (LC) has been implicated in a variety of physiological functions including sleep/wakefulness, cognition/memory, stress/emotion, and pain. Marked loss of LC-noradrenergic (NAergic) neurons is observed in autopsy specimens of patients with Alzheimer's disease and Parkinson's disease (PD), and part of the clinical symptoms of these diseases may be related to dysfunction of the LC. Neurotoxins have been utilized to ablate LC-NAergic neurons in experimental animals for elucidating the pathophysiological implication of the loss of LC, but there are methodological drawbacks in previously utilized methods. We employed immunotoxin-mediated neuronal targeting to overcome these problems. Following complete disruption of the LC-NAergic neurons by immunotoxin, mice showed behavioral changes, which resembled the nonmotor symptoms of PD. The LC-NAergic neurons did not regenerate following ablation, so the immunotoxin-mediated neuronal targeting may be useful especially for studying the long-term effects of the loss of LC-NAergic neurons on brain functions.
 
Article
Mitochondria from persons with Alzheimer's disease (AD) differ from those of age-matched control subjects. Differences in mitochondrial morphology and function are well documented, and are not brain-limited. Some of these differences are present during all stages of AD, and are even seen in individuals who are without AD symptoms and signs but who have an increased risk of developing AD. This chapter considers the status of mitochondria in AD subjects, the potential basis for AD subject mitochondrial perturbations, and the implications of these perturbations. Data from multiple lines of investigation, including epidemiologic, biochemical, molecular, and cytoplasmic hybrid studies, are reviewed. The possibility that mitochondria could potentially constitute a reasonable AD therapeutic target is discussed, as are several potential mitochondrial medicine treatment strategies.
 
Article
A genetic defect in habituation in schizophrenic patients results from malfunction in the limbic system of the brain, which is correctable by administration of propranolol but not by phenothiazines. It is proposed in the chapter that the disturbances in communication patterns among subunits of the limbic system that are largely coordinated by activities of the GABAergic system result from mutation of the SCN5A gene, which uniquely codes for the major voltage‐gated Na+-channel in the limbic system. Therapeutic problems in large‐scale clinical use of propranolol may arise because the SCN5A gene also codes for the major Na+ channel in the heart; and appropriate use of propranolol requires maintenance of a balance between achieving behavioral improvement and occurrence of cardiac insufficiency. Conditions for use of propranolol or similarly effective substances are being sought that maximize the former and minimize the latter.
 
Article
The ubiquitin-proteasome complex is an important molecular target for the design of novel chemotherapeutics. This complex plays a critical role in signal transduction pathways important for tumor cell growth and survival, cell-cycle control, transcriptional regulation, and the modulation of cellular stress responses to endogenous and exogenous stimuli. The sensitivity of transformed cells to proteasome inhibitors and the successful design of treatment protocols with tolerable, albeit narrow, therapeutic indices have made proteasome inhibition a viable strategy for cancer treatment. Clinical validation of the proteasome as a molecular target was achieved with the approval of bortezomib, a boronic acid proteasome inhibitor, for the treatment of multiple myeloma and mantle cell lymphoma. Several "next-generation" proteasome inhibitors (carfilzomib and PR-047, NPI-0052, and CEP-18770) representing distinct structural classes (peptidyl epoxyketones, beta-lactones, and peptidyl boronic acids, respectively), mechanisms of action, pharmacological and pharmacodynamic activity profiles, and therapeutic indices have now entered clinical development. These agents may expand the clinical utility of proteasome inhibitors for the treatment of solid tumors and for specific non-oncological, i.e., inflammatory disease, indications as well. This chapter addresses the biology of the proteasome, the medicinal chemistry and mechanisms of action of proteasome inhibitors currently in clinical development, the preclinical and clinical pharmacological and safety profiles of bortezomib and the newer compounds against hematological and solid tumors. Future directions for research and other applications for this novel class of therapeutics agents are considered in this chapter.
 
Article
Monoclonal antibody-based medications designed to bind (+)-methamphetamine (METH) with high affinity are among the newest approaches to the treatment of METH abuse and the associated medical complications. The potential clinical indications for these medications include treatment of overdose, reduction of drug dependence, and protection of vulnerable populations from METH-related complications. Research designed to discover and conduct preclinical and clinical testing of these antibodies suggests a scientific vision for how intact monoclonal antibody (mAb) (singular and plural) or small antigen-binding fragments of mAb could be engineered to optimize the proteins for specific therapeutic applications. In this review, we discuss keys to success in this development process including choosing predictors of specificity, efficacy, duration of action, and safety of the medications in disease models of acute and chronic drug abuse. We consider important aspects of METH-like hapten design and how hapten structural features influence specificity and affinity, with an example of a high-resolution X-ray crystal structure of a high-affinity antibody to demonstrate this structural relationship. Additionally, several prototype anti-METH mAb forms such as antigen-binding fragments and single-chain variable fragments are under development. Unique, customizable aspects of these fragments are presented with specific possible clinical indications. Finally, we discuss clinical trial progress of the first in kind anti-METH mAb, for which METH is the disease target instead of vulnerable central nervous system networks of receptors, binding sites, and neuronal connections.
 
Effects of 7-day treatment with saline or transporter substrates on responding maintained by cocaine (0.01 mg/kg/injection) or food pellets by rhesus monkeys. Monkeys could earn a maximum of 80 cocaine injections and 100 food pellets each day during alternating components of cocaine and food availability. Treatment consisted of continuous infusion with saline (upper left) or optimal treatment doses of the DAT/NET>SERT-selective substrate methamphetamine (0.056 mg/kg/hr), the nonselective DAT/NET/SERT substrate PAL-287 (1.0 mg/kg/hr) or the SER>DAT/NET-selective substrate fenfluramine (1.0 mg/kg/hr). All substrates produced sustained decreases in cocaine self-administration, and behavioral selectivity to decrease cocaine-maintained responding corresponded to pharmacological selectivity for DAT/NET vs. SERT. Adapted from Negus et al. 2007.
Article
The acute and chronic effects of abused psychostimulants on monoamine transporters and associated neurobiology have encouraged development of candidate medications that target these transporters. Monoamine transporters, in general, and dopamine transporters, in particular, are critical molecular targets that mediate abuse-related effects of psychostimulants such as cocaine and amphetamine. Moreover, chronic administration of psychostimulants can cause enduring changes in neurobiology reflected in dysregulation of monoamine neurochemistry and behavior. The current review will evaluate evidence for the efficacy of monoamine transporter inhibitors and substrates to reduce abuse-related effects of stimulants in preclinical assays of stimulant self-administration, drug discrimination, and reinstatement. In considering deployment of monoamine transport inhibitors and substrates as agonist-type medications to treat stimulant abuse, the safety and abuse liability of the medications are an obvious concern, and this will also be addressed. Future directions in drug discovery should identify novel medications that retain efficacy to decrease stimulant use but possess lower abuse liability and evaluate the degree to which efficacious medications can attenuate or reverse neurobiological effects of chronic stimulant use.
 
Article
Acute activation of kappa-opioid receptors produces anti-addictive effects by regulating dopamine levels in the brain. Unfortunately, classic kappa-opioid agonists have undesired side effects such as sedation, aversion, and depression, which restrict their clinical use. Salvinorin A (Sal A), a novel kappa-opioid receptor agonist extracted from the plant Salvia divinorum, has been identified as a potential therapy for drug abuse and addiction. Here, we review the preclinical effects of Sal A in comparison with traditional kappa-opioid agonists and several new analogs. Sal A retains the anti-addictive properties of traditional kappa-opioid receptor agonists with several improvements including reduced side effects. However, the rapid metabolism of Sal A makes it undesirable for clinical development. In an effort to improve the pharmacokinetics and tolerability of this compound, kappa-opioid receptor agonists based on the structure of Sal A have been synthesized. While work in this field is still in progress, several analogs with improved pharmacokinetic profiles have been shown to have anti-addictive effects. While in its infancy, it is clear that these compounds hold promise for the future development of anti-addictive therapeutics.
 
Article
Drug abuse continues to create a major international epidemic affecting society. A great majority of past drug abuse research has focused mostly on the mechanisms of addiction and the specific effects of substance use disorders on brain circuits and pathways that modulate reward, motivation, craving, and decision making. Few studies have focused on the neurobiology of acute and chronic substance abuse as it relates to the neurovascular unit (brain endothelial cell, neuron, astrocyte, microglia, and pericyte). Increasing research indicates that all cellular components of the neurovascular unit play a pivotal role in both the process of addiction and how drug abuse affects the brain response to diseases. This review will focus on the specific effects of opioids, amphetamines, alcohol, and nicotine on the neurovascular unit and its role in addiction and adaption to brain diseases. Elucidation of the role of the neurovascular unit on the neurobiology associated with drug addiction will help to facilitate the development of better therapeutic approaches for drug-dependent individuals.
 
Article
Drugs of abuse and Fonzies (palatable food) share the properties of stimulating dopamine (DA) transmission within the nucleus accumbens “shell,” as compared with the “core” Nonpsychostimulant drugs of abuse and food depend for their property of stimulating DA release in the shell of the nucleus accumbens on an activation of the μ1 subtype of opioid receptors. Substances of abuse and Fonzies show differences in their ability to influence DA transmission in the prefrontal cortex. Fonzies activate DA release in the prefrontal cortex, but drugs of abuse, as a whole, do not activate DA transmission in the prefrontal cortex. Studies on Fonzies show that DA plays a different role in the prefrontal cortex and in the nucleus accumbens. It appears that while prefrontal DA transmission reacts to generically salient stimuli independently of motivational state, nucleus accumbens DA reacts to these stimuli only under a deprivation state. Under a nondeprivation state, nucleus accumbens DA reacts only to strong, unconditioned, novel, a d unpredicted stimuli. It is suggested that phasic stimulation of DA release in the shell of the nucleus accumbens by a conventional reinforcer such as food is not essential for the expression of incentive behavior and for the hedonic sensations arising from consumption of the reward. Thus, naloxonazine, a slowly reversible μ1 antagonist, blocks the stimulation of DA release in the nucleus accumbens shell by morphine, nicotine, and ethanol, as well as by palatable food. Therefore, under nondeprivation conditions, stimulation of dopamine release in the shell of the nucleus accumbens might be related to the acquisition rather than the expression of motivated behavior.
 
Article
D1-receptor antagonists block dopamine agonist-induced gene expression in striatonigral neurons. However, D2 antagonists also significantly attenuate these effects, contributing to the idea that both D1 and D2 receptors must be stimulated to evoke gene expression in striatonigral neurons. Although D1 receptors are thought to control striatonigral gene expression, several studies have demonstrated that D2-receptor antagonists block stimulantinduced dynorphin immunoreactivity and preprodynorphin (PPD) gene expression. In addition to the strong regulation of gene expression in medium spiny neurons by dopamine receptors, recent studies indicate that glutamatergic and cholinergic neurotransmission strongly modulates the effects of dopamine on striatal gene expression. Glutamatergic regulation of the alterations in expression of immediate early genes (IEGs) and neuropeptide mRNAs and their protein levels in the striatum may be essential to the neuronal adaptation that underlies delayed and long-term effects of psychostimulants. Stimulant-induced IEG and ne ropeptide expression in striatal neurons has been linked both to N-methyl-D-aspartate (NMDA) and non-NMDA receptor activation. Apart from this, acetylcholine also regulates striatal gene expression. Acetylcholine and dopamine also exert opposite effects on striatopallidal gene expression. At the level of the substantia nigra, D1-receptor stimulation that facilitates local GABA release may result in disinhibition of glutamate release in the thalamostriatal and corticostriatal pathways and further stimulation of striatal acetylcholine release. The inhibitory influence of cholinergic interneurons on striatonigral neurons also provides an intrastriatal mechanism for some D1/D2 interactions. Thus, both muscarinic receptor agonists and excitatory amino acid receptor antagonists are potentially useful tools in the search for novel therapeutic approaches to drug abuse.
 
Article
Despite the proven efficacy of current pharmacotherapies for tobacco dependence, relapse rates continue to be high, indicating that novel medications are needed. Currently, several smoking cessation agents are available, including varenicline (Chantix®), bupropion (Zyban®), and cytisine (Tabex®). Varenicline and cytisine are partial agonists at the α4β2* nicotinic acetylcholine receptor (nAChR). Bupropion is an antidepressant but is also an antagonist at α3β2* ganglionic nAChRs. The rewarding effects of nicotine are mediated, in part, by nicotine-evoked dopamine (DA) release leading to sensitization, which is associated with repeated nicotine administration and nicotine addiction. Receptor antagonists that selectivity target central nAChR subtypes mediating nicotine-evoked DA release should have efficacy as tobacco use cessation agents with the therapeutic advantage of a limited side-effect profile. While α-conotoxin MII (α-CtxMII)-insensitive nAChRs (e.g., α4β2*) contribute to nicotine-evoked DA release, these nAChRs are widely distributed in the brain, and inhibition of these receptors may lead to nonselective and untoward effects. In contrast, α-CtxMII-sensitive nAChRs mediating nicotine-evoked DA release offer an advantage as targets for smoking cessation, due to their more restricted localization primarily to dopaminergic neurons. Small drug-like molecules that are selective antagonists at α-CtxMII-sensitive nAChR subtypes that contain α6 and β2 subunits have now been identified. Early research identified a variety of quaternary ammonium analogs that were potent and selective antagonists at nAChRs mediating nicotine-evoked DA release. More recent data have shown that novel, nonquaternary bis-1,2,5,6-tetrahydropyridine analogs potently inhibit (IC50<1nM) nicotine-evoked DA release in vitro by acting as antagonists at α-CtxMII-sensitive nAChR subtypes; these compounds also decrease NIC self-administration in rats.
 
Article
Methamphetamine abuse escalates, but no approved therapeutics are available to treat addicted individuals. Methamphetamine increases extracellular dopamine in reward-relevant pathways by interacting at vesicular monoamine transporter-2 (VMAT2) to inhibit dopamine uptake and promote dopamine release from synaptic vesicles, increasing cytosolic dopamine available for reverse transport by the dopamine transporter (DAT). VMAT2 is the target of our iterative drug discovery efforts to identify pharmacotherapeutics for methamphetamine addiction. Lobeline, the major alkaloid in Lobelia inflata, potently inhibited VMAT2, methamphetamine-evoked striatal dopamine release, and methamphetamine self-administration in rats but exhibited high affinity for nicotinic acetylcholine receptors (nAChRs). Defunctionalized, unsaturated lobeline analog, meso-transdiene (MTD), exhibited lobeline-like in vitro pharmacology, lacked nAChR affinity, but exhibited high affinity for DAT, suggesting potential abuse liability. The 2,4-dicholorophenyl MTD analog, UKMH-106, exhibited selectivity for VMAT2 over DAT, inhibited methamphetamine-evoked dopamine release, but required a difficult synthetic approach. Lobelane, a saturated, defunctionalized lobeline analog, inhibited the neurochemical and behavioral effects of methamphetamine; tolerance developed to the lobelane-induced decrease in methamphetamine self-administration. Improved drug-likeness was afforded by the incorporation of a chiral N-1,2-dihydroxypropyl moiety into lobelane to afford GZ-793A, which inhibited the neurochemical and behavioral effects of methamphetamine, without tolerance. From a series of 2,5-disubstituted pyrrolidine analogs, AV-2-192 emerged as a lead, exhibiting high affinity for VMAT2 and inhibiting methamphetamine-evoked dopamine release. Current results support the hypothesis that potent, selective VMAT2 inhibitors provide the requisite preclinical behavioral profile for evaluation as pharmacotherapeutics for methamphetamine abuse and emphasize selectivity for VMAT2 relative to DAT as a criterion for reducing abuse liability of the therapeutic.
 
Article
This chapter reviews several recent cases in which transgenic mouse and human data provide increasingly compelling evidence that variants in genes important for dopaminergic neurotransmission are strong candidates to contribute to interindividual differences in drug abuse vulnerabilities. Studies of cocaine's primary site for reward and reinforcement in the brain have focused on the dopamine transporter (DAT). In studying this transporter, the effects of several mutations on transporter function have been characterized. To model the effects that a DAT gene variant might have on drug abuse phenotypes, the researchers have constructed transgenic animals that express this DAT variant in catecholaminergic neurons. In these animals, although baseline locomotor activities are altered, cocaine induces virtually no excess locomotion. Vesicular monoamine stores accumulated by normal vesicular monoamine transporter (VMAT2) function may play significant roles in the locomotor stimulation and/or the behavioral reward produced by amphetamines. Catechol-O-Methyltransferase (COMT) is expressed in dopaminergic brain regions, where its activity provides a major pathway by which extraneuronally released dopamine is inactivated. Significant differences in the distributions of both COMT genotypes and allele frequencies between controls and substance abusers have been identified. The data suggest that high COMT activities might contribute to the genetic underpinnings of drug abuse vulnerability. Thus the chapter supports potentially prominent roles for dopaminergic gene variants in contributing to individual differences in vulnerability to drug abuse. However, other genes are quite likely also to be involved.
 
Panel A: Results on distance travelled (cm) by mice treated b.i.d. for seven days with ibudilast (IBUD) or its vehicle (VEH1), beginning two days before five days of treatment with 3 mg/kg methamphetamine. Ibudilast was administered at 1.8, 7.5, or 13 mg/kg. Data points represent group means (±S.E.M.) obtained during 1-h experimental sessions. Filled data points represent sessions preceded by 3 mg/kg i.p. methamphetamine injections. Unfilled data points represent sessions preceded by i.p. saline injections. N=8 for each treatment group. *P<0.05 with respect to mice treated with ibudilast's vehicle. Modified and reprinted with permission from reference (Snider et al., 2012). Panel B: Effects of ibudilast or its vehicle on group mean infusions of methamphetamine (0.001, 0.03, and 0.1 mg/kg/inf) obtained during daily 2-h self-administration sessions. Ibudilast was administered at 1, 7.5, or 10 mg/kg i.p. b.i.d. for 3 consecutive days at each methamphetamine self-administered dose. Data points represent the group means of total infusions obtained during the third day of testing at each ibudilast dose. Bars through symbols indicate ±S.E.M. Data point above "S" on the abscissa indicates results when saline
Article
Glia (including astrocytes, microglia, and oligodendrocytes), which constitute the majority of cells in the brain, have many of the same receptors as neurons, secrete neurotransmitters and neurotrophic and neuroinflammatory factors, control clearance of neurotransmitters from synaptic clefts, and are intimately involved in synaptic plasticity. Despite their prevalence and spectrum of functions, appreciation of their potential general importance has been elusive since their identification in the mid-1800s, and only relatively recently have they been gaining their due respect. This development of appreciation has been nurtured by the growing awareness that drugs of abuse, including the psychostimulants, affect glial activity, and glial activity, in turn, has been found to modulate the effects of the psychostimulants. This developing awareness has begun to illuminate novel pharmacotherapeutic targets for treating psychostimulant abuse, for which targeting more conventional neuronal targets has not yet resulted in a single, approved medication. In this chapter, we discuss the molecular pharmacology, physiology, and functional relationships that the glia have especially in the light in which they present themselves as targets for pharmacotherapeutics intended to treat psychostimulant abuse disorders. We then review a cross section of preclinical studies that have manipulated glial processes whose behavioral effects have been supportive of considering the glia as drug targets for psychostimulant-abuse medications. We then close with comments regarding the current clinical evaluation of relevant compounds for treating psychostimulant abuse, as well as the likelihood of future prospects.
 
Article
Drugs of abuse can be divided, on the basis of their electrophysiological effects, into two classes: psychostimulants that decrease dopaminergic activity, and others, the acute administration of which increases dopamine firing rate and pattern. Chronic administration of drugs may mimic better neuronal alterations relevant for understanding mechanisms of drug dependence. The mesolimbic dopamine system is a major target of drugs of abuse, not only after acute administration, but also after a chronic challenge. While the chronic intake of drugs of abuse represents an essential step in the development of drug addiction, the withdrawal syndrome is generally the painful end. The withdrawal syndrome is, thus, an extremely important phase in the process of drug dependence because it combines the positive and negative reinforcing properties of drugs of abuse. This chapter summarizes a series of experiments aimed at investigating the physiological status of mesolimbic dopaminergic neurons during and after withdrawal from two of the most abused drugs: ethanol and morphine. The studies lead us to the conclusion that the apparent depolarization block requires an essential ingredient such as the use of chloral hydrate and thus, is extremely unlikely to be the effect of ethanol withdrawal. Similarly, dopamine neurons recorded from rats made dependent on morphine while experiencing a measurable withdrawal syndrome showed electrophysiological features reminiscent of those observed in ethanolwithdrawn rats. The striking similarity of results obtained with two different chemicals, such as ethanol and morphine, reinforces the view that the mesolimbic dopamine system is profoundly affected by chronic administration and withdrawal of addicting drugs and consequently may be viewed as the target for potential new therapies aimed at ameliorating the psychological discomfort produced by drug withdrawal.
 
Article
A drug with slower onset of action would have less abuse liability. In principle, these two favorable characteristics could be achieved either pharmacokinetically, such as by giving the drug via a slowly absorbed route of administration or pharmacodynamically, such as by giving an analogue with slow, long-lasting binding to the site of action. Evidence in humans for the rate hypothesis of cocaine effects is largely circumstantial, based on pharmacokinetic differences among routes of administration. The agents with slow onset of effect, including possibly the abused drug itself by another route of administration, may be useful in implementing the agonist-substitution approach to treatment of cocaine abuse, consistent with the rate hypothesis of psychoactive drug effect. In this chapter several medications that bind to the presumed major site of action for cocaine's psychoactive effects have been studied as possible agonist substitution agents. These agents have some mild stimulant-like agonist properties, with little apparent human abuse potential when taken in slow-onset form. Both mazindol, an appetite suppressant, and bupropion, an antidepressant, have been ineffective in double-blind clinical trials. This may be related to dose-limiting side effects that have kept doses in the range that probably occupies less than half of brain presynaptic dopamine transporters (DAT) sites. In animal studies, it has reduced the increased brain extracellular dopamine concentration produced by cocaine administration and reduced cocaine selfadministration at doses that do not influence food intake.
 
Article
Neuroadaptations within the nucleus accumbens (NAc) appear to be most responsible for the expression of many indices of cocaine withdrawal, including behavioral sensitization. In this chapter a close temporal relationship between the persistence of behavioral sensitization and enhanced responsiveness of NAc neurons to DA D1-receptor stimulation has been demonstrated. Cocaine-pretreated NAc neurons exhibit a number of significant alterations in passive and active membrane properties, as compared with saline-pretreated controls. Depolarization of NAc neurons by intracellular current injection revealed additional significant differences between cocaine-pretreated and control neurons. Accordingly, neuromodulation of PKC activity might also be altered by repeated cocaine administration in such a way as to enhance basal states of phosphorylation. Several of the effects produced by repeated cocaine administration, in particular the increased threshold, decreased action potential amplitude, and reduction in repetitive firing, might be indicative of alterations in voltagesensitive sodium channels (VSSCs). The reduction in peak Na+ current produced by repeated cocaine treatment is consistent with enhanced phosphorylation of VSSCs. The level of VSSC activity is subject to tonic modulation by the levels of protein kinase (PKA), an enzyme that is known to be increased within the NAc by repeated cocaine administration. Although alterations in transmission at specific synapses are certain to modify the responsiveness of NAc neurons to selected inputs, the marked reduction of VSSC function can produce a more profound and indiscriminate decrease in the responsiveness of the NAc to exitatory commands.
 
Article
Vesicular transporters of transmitters are excellent markers for functional neuroanatomy and for potential identification of vesicular contents at the subcellular level. In this chapter, the distribution of two vesicular monoamine transporters (VMAT1 and VMAT2), a vesicular acetylcholine transporter (VAChT), a transmembrane transporter (SV2), and two other proteins have been examined. VMAT1 is endogenous in PC12 cells, and it is abundant on LDCVs. This location of VMAT1 is consistent with the fact that LDCVs take up norepinephrine in PC12 cells. On the otherhand, VMAT2 is not endogenous in PC12 cells. In the chapter VMAT2 has been expressed in PC12 cells under a constitutive viral promoter to see if it would be sorted to secretory vesicles. In the transfected PC12 cells, VMAT2 is preferentially localized on the LDCVs and not on the SSVs. The lack of a prominent population of VMAT2-positive synaptic vesicles in these PC12 cells is in contrast to the distribution of endogenous VAMT2 in central and peripheral adrenergic neurons. In these adrenergic neurons in vivo, VMAT2 is localized on LDCVs and on a distinct population of catecholamine-containing synaptic vesicles that are termed small dense-core vesicles (SDCVs). The chapter also analyzes subcellular distributions of three other SSV components, synaptophysin, SV2, and VAChT. Proteins destined for cholinergic SSVs may arrive via LDCVs or constitutive vesicles and recycle through the early endosomes, where they are concentrated for inclusion in the SSV membrane. Proteins in this category include SV2, VAChT, and synaptophysin. VMAT2, the neuronal VMAT in LDCVs and SDCVs of noradrenergic neurons, is not targeted to cholinergic SSVs when expressed in PC12 cells. Thus, SDCVs of noradrenergic neurons may represent a distinct population of synaptic vesicles that is biosynthetically separate from cholinergic SSVs in PC12 cells.
 
Article
The search for the physiological function of nicotinic receptors on neurons in the brain began with their discovery. It was initially assumed that, as in ganglia and at the neuromuscular junction, nicotinic receptors would gate fast synaptic transmission in the brain. The best functional evidence now, however, points to a role in modifying the release of other transmitters. This does not preclude a postsynaptic role in transmission for nicotinic receptors in the brain, but attempts to locate such a synapse have not been successful. If fast nicotinic synapses are present in the brain, they are probably low in number and may be masked by other more prevalent synapses (such as glutamatergic) so identification will not be easy. The extent of diversity of nicotinic receptors is substantial. At the molecular level this is reflected in the number of different genes that encode receptor subunits and the multiple possible combinations of subunits that function in expression systems. From the cellular level there is a broad diversity of properties of native receptors in neurons. Some useful pharmacological tools allow the limited identification of subunits in native receptors. For example, block by alpha-bungarotoxin identifies alpha 7, alpha 8, or alpha 9 subunits; activation of a receptor by cytisine indicates an alpha 7 or beta 4 subunit; and neuronal bungarotoxin block identifies a beta 2 subunit. Despite the clues to identity gained by careful use of these agents, we have not been able to identify all the components of any native receptor based on pharmacological properties assessed from expression studies. When both pharmacological and biophysical properties of a receptor are taken into consideration, none of the combinations tested in oocytes mimics native receptors exactly. The reason for this discrepancy has been debated at length; it is possible that oocytes do not faithfully manufacture neuronal nicotinic receptors. For example, they may not correctly modify the protein after translation or they may allow a combination of subunits that do not occur in vivo. Another possibility is that correct combinations of subunits have not yet been tested in oocytes. Data from immunoprecipitation experiments suggest that many receptors contain three or more different subunits. Results from further experiments injecting combinations of three or more subunits into oocytes may be enlightening. The diversity of receptors may allow targeting of subtypes to specific locations. Nicotinic receptors are located presynaptically, preterminally, and on the cell soma. The function of the nicotinic receptors located on innervating axons is presumably to modify the release of other neurotransmitters. It is an attractive hypothesis that nicotinic receptors might be involved in modifying the weight of central synapses; however, in none of the regions where this phenomenon has been described is there any evidence for axoaxonal contacts. The presynaptic receptors described so far are pharmacologically unique; therefore, if there are different subtypes of nicotinic receptors modifying the release of different transmitters, they may provide a means of exogenously modifying the release of a particular transmitter with drugs. There are still many basic unanswered questions about nicotinic receptors in the brain. What are the compositions of native nicotinic receptors? What is their purpose on neurons? Although there is clearly a role presynaptically, what is the function of those located on the soma? Neuronal nicotinic receptors are highly permeable to calcium, unlike muscle nicotinic receptors, and this may have important implications for roles in synaptic plasticity and development. Finally, why is there such diversity? (ABSTRACT TRANCATED)
 
Article
The diversity of application of the thiol drug NAC in both the experimental setting, as a tool for the study of the mechanisms and consequences of oxidative stress, and the clinical setting, as a therapeutic agent, clearly reflects the central role played by the redox chemistries of the group XVI elements, oxygen and sulfur, in biology. As our understanding of such redox processes increases, particularly their roles in specific pathophysiological processes, new avenues will open for the use of NAC in the clinical setting. As a drug, NAC represents perhaps the ideal xenobiotic, capable of directly entering endogenous biochemical processes as a result of its own metabolism. Thus, it is hoped that the experience gained with this unique agent will help in future efforts to design antioxidants and chemoprotective principles which are able to more accurately utilize endogenous biochemical processes for cell- or tissue-specific therapy.
 
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Massive loss of skeletal muscle mass (cachexia) is sign of HIV infection, sepsis, and trauma, and a cause of death among cancer patients. In HIV infection, the immunological dysfunction develops progressively into a severe condition called acquired immunodeficiency syndrome. Increasing evidence suggests that an abnormal cysteine and glutathione metabolism plays a decisive role in the development of catabolic conditions and associated immunological dysfunctions. This chapter gives a conceptual overview; the chances for a therapeutic intervention with cysteine derivatives such as N-acetylcysteine (NAC) and the methodological aspects. The agenda is to identify abnormal biochemical patterns in different catabolic diseases and conditions, abnormal biochemical parameters that are significantly correlated with mortality, disease progression, and weight loss, to prove cause-and-effect relationships by experimental intervention, to develop and optimize interventive strategies that may be suitable for clinical therapy, and to test new therapeutic strategies in clinical trials. Discussed are the elevated venous plasma glutamate levels as universal marker for catabolic and precatabolic conditions—evidences for an impairment of muscular membrane transport activities, abnormal cysteine catabolism and glutathione level in skeletal muscle tissue, and the pathological significance of decreased membrane transport activity and elevated venous plasma glutamate levels. There is discussion on “Push” and “Pull” mechanisms in catabolic and precatabolic processes, decreased plasma cystine levels and evidence for “Push” and “Pull” mechanisms in sepsis, HIV Infection, and other malignant diseases, abnormally low plasma cystine and glutamine levels in patients with chronic fatigue syndrome (CFS), and the hypothetical mechanism of the “pull” condition—evidence for an abnormal cysteine and glutathione metabolism in liver. Noted are the immunological implications—cysteine deficiency and immunological dysfunction, abnormal glutathione levels in HIV Infection, and redox regulation by glutathione and glutathione disulfide. Also, there are details on the therapeutic intervention with a cysteine derivative: effects of long-term treatment with N-acetylcysteine.
 
Article
Arylamine N-acetyltransferases (NATs) are defined as xenobiotic metabolizing enzymes, adding an acetyl group from acetyl coenzyme A (CoA) to arylamines and arylhydrazines. NATs are found in organisms from bacteria and fungi to vertebrates. Several isoenzymes, often polymorphic, may be present in one organism. There are two functional polymorphic NATs in humans and polymorphisms in NAT2 underpinned pharmacogenetics as a discipline. NAT enzymes have had a role in important metabolic concepts: the identification of acetyl-CoA and endogenous metabolic roles in bacteria and in eukaryotic folate metabolism. In fungi, NAT is linked to formation of unique metabolites. A broad and exciting canvas of investigations has emerged over the past five years from fundamental studies on NAT enzymes. The role of human NAT1 in breast cancer where it is a biomarker and possible therapeutic target may also underlie NAT's early appearance during mammalian fetal development. Studies of NAT in Mycobacterium tuberculosis have identified potential therapeutic targets for tuberculosis whilst the role of NATs in fungi opens up potential toxicological intervention in agriculture. These developments are possible through the combination of genomics, enzymology and structural data. Strong binding of CoA to Bacillis anthracis NAT may point to divergent roles of NATs amongst organisms as does differential control of mammalian NAT gene expression. The powerful combination of phenotypic investigation following genetic manipulation of NAT genes from mice to mycobacteria has been coupled with generation of isoenzyme-specific inhibitors. This battery of molecular and systems biology approaches heralds a new era for NAT research in pharmacology and toxicology.
 
Article
Acetyltransferases play a central role in the metabolic disposition, detoxication, and bioactivation of a diverse group of drugs—carcinogens and other xenobiotics. Acetylation is a major metabolic pathway for primary aromatic amines (arylamines, ArNH) and hydrazines. In earlier times, acetylation was the principal mechanism for termination of the tuberculostatic action of isoniazid. The importance of human acetylation capacity is further illustrated by the propensity of phenotypically slow acetylator patients to experience methemoglobinemia or drug-induced lupus erythematosus or both when treated either with arylamine drugs or with various agents that contain the hydrazine moiety. The mutagenicity and carcinogenicity of arylamines along with thedocumented environmental and dietary exposure to these agents have stimulated numerous investigations of arylamine metabolism. Thus, the premise that formation of unreactive arylamides by N-acetylation of arylamines is primarily a detoxification process and that oxidative Nhydroxylation is a toxification reaction is supported by a substantial body of evidence. However, the demonstration that arylamine N-acetyltransferases are versatile enzymes that can catalyze the conversion of both N-arylhydroxylamines and N-arylhydroxamic acids to reactive, electrophilic metabolites has broadened the scope of research on acetyltransferases, which are now included among those conjugation systems that play important roles in the bioactivation of xenobiotics.
 
Article
Metabolic acidosis is common during cardiovascular dysfunction. The effect of acidosis on cardiovascular function has been extensively studied. This chapter summarizes several studies that evaluate the cardiovascular effects of acute infusion of lactic acid or hydrochloric acid and compared the cardiovascular effects of dobutamine (DOB) and epinephrine (EPI) as well as norepinephrine (NE) on the cardiovascular system. In the present study there has been, in general, a decrease in the responsiveness to the infusion of endogenous catecholamines in acidotic animals compared to subjects with normal pH. Epinephrine remains the drug of choice for cardiac arrest and severe, refractory hypotension, and it has been suggested that the most effective dose may be higher than those currently recommended during cardiopulmonary resuscitation (CPR). There are several possible mechanisms for the deterioring cardiovascular responses to catecholamine stimulation. First, myocardial intracellular acidosis from accumulation of lactic acidosis (LAC) may contribute directly to the impairment of myocardial function. Second, the prolonged sympatho-adrenal stimulation that results from acidosis may result in depletion of endogenous catecholamines and energy substances and progressive deterioration of myocardial contractility. Third, the pulmonary and systemic vasoconstriction induced by acidosis and the infusions of EPI or NE will increase afterload to the myocardium. Although the response to endogenous catecholamines is depressed in the acidotic animal, a more normal dose-response relationship with DOB on the cardiovascular parameters has been measured. There are reports that acidosis created by LAC infusion produces hypoxic-induced increases in pulmonary arterial pressure or pulmonary vascular resistance. The increase in pulmonary vascular resistance caused by the infusion of HCl may be mediated by thromboxane A2 or prostacyclin synthesis.
 
Article
Epoxyeicosatrienoic acids (EETs) are cytochrome P450 metabolites of arachidonic acid that are produced by the vascular endothelium in responses to various stimuli such as the agonists acetylcholine (ACH) or bradykinin or by shear stress which activates phospholipase A(2) to release arachidonic acid. EETs are important regulators of vascular tone and homeostasis. In the modulation of vascular tone, EETs function as endothelium-derived hyperpolarizing factors (EDHFs). In models of vascular inflammation, EETs attenuate inflammatory signaling pathways in both the endothelium and vascular smooth muscle. Likewise, EETs regulate blood vessel formation or angiogenesis by mechanisms that are still not completely understood. Soluble epoxide hydrolase (sEH) converts EETs to dihydroxyeicosatrienoic acids (DHETs) and this metabolism limits many of the biological actions of EETs. The recent development of inhibitors of sEH provides an emerging target for pharmacological manipulation of EETs. Additionally, EETs may initiate their biological effects by interacting with a cell surface protein that is a G protein-coupled receptor (GPCR). Since GPCRs represent a common target of most drugs, further characterization of the EET receptor and synthesis of specific EET agonists and antagonist can be used to exploit many of the beneficial effects of EETs in vascular diseases, such as hypertension and atherosclerosis. This review will focus on the current understanding of the contribution of EETs to the regulation of vascular tone, inflammation, and angiogenesis. Furthermore, the therapeutic potential of targeting the EET pathway in vascular disease will be highlighted.
 
Article
This chapter summarizes various studies to extend published findings in skeletal muscle regarding the actions of fatty acids. In the absence of anesthetics, the fatty acids can cause Ca2+ release in skeletal muscle and rabbit cardiac muscle by mechanisms independent of the Ca2+ release channel. Similar interactions could occur with other proteins, such as the Na+ channel, that also has its function altered by halothane and fatty acids. However, significant interactions of free fatty acids with the Ca2+ release channel and Na+ channel are somewhat unlikely under normal circumstances in an intact cell, as the fatty acids are bound to fatty acid-binding proteins in the cytoplasm. Although fatty acids probably have very little impact on Ca2+ regulation in the absence of anesthetics, their augmentation of net halothane-induced Ca2+ release is dramatic. Whereas the concentration of fatty acid required for this effect exceeds that of the normally unbound form, halothane could likely displace fatty acids from fatty acid-binding proteins. Considering the low concentration of fatty acids present and the resistance of the cardiac preparation to fatty acid-albumin complexes, it is unlikely that displacement of fatty acids by halothane plays a significant role in anesthetic action in rat cardiac muscle. Human or porcine skeletal muscle contains about 5 to 10 times as much free fatty acid as cardiac muscle. The huge potency of fatty acid-albumin complexes in their interaction with halothane in skeletal muscle compared to rat cardiac muscle suggests some actual association of fatty acid-binding proteins with the ryanodine receptor.
 
Top-cited authors
Anthony Cerami
  • Araim Pharmaceuticals
Richard Bucala
  • Yale University
Ziang Yan
  • Tsinghua University
Dalton James Surmeier
  • Northwestern University
Wen-Jie Song
  • Kumamoto University