Donard S. Dwyer

Induction of hyperglycemia in mice with atypical antipsychotic drugs that inhibit glucose uptake.

Many antipsychotic drugs disturb the regulation of glucose metabolism in patients treated for schizophrenia. The goal of the present studies was to determine if these antipsychotic drugs produce hyperglycemia in mice in relation to their ability to interfere with glucose uptake and utilization. Male C57BL/6 mice were injected with a panel of typical and atypical antipsychotic drugs and blood glucose levels were determined periodically over a 3- to 6-h time interval. The atypical drugs, clozapine, desmethylclozapine, quetiapine, and loxapine, and the original antipsychotic, chlorpromazine, induced significant hyperglycemia in the mice in accordance with their effects on glucose transport. By contrast, haloperidol and sulpiride, which have little effect on glucose uptake, did not induce hyperglycemia. Risperidone produced a modest elevation of blood glucose levels, but only at a low dose of the drug. Cytochalasin B, a specific inhibitor of the glucose transporter (GLUT) protein, produced significant hyperglycemia in the mice. Overall, there was a strong correlation between the ability of a drug to inhibit glucose transport in vitro and its ability to induce hyperglycemia in vivo. Finally, the drugs that produced hyperglycemia in mice have been linked to the development of diabetes in patients.

Antipsychotic drugs affect glucose uptake and the expression of glucose transporters in PC12 cells.

1. Adherence of the PC12 cell line to poly-l-lysine (PLL) on tissue culture dishes stimulated glucose transport into the cells. Fluphenazine, chlorpromazine, clozapine and haloperidol inhibited glucose uptake in this system after a short (30 min) preincubation with drug. The IC50s for this effect were typically in the range of 5-40 microM. 2. Following longer exposures of the drugs (24 hr), there was a significant increase (approximately 3-fold) in the cellular levels of the glucose transporter (GLUT) isoforms, GLUT1 and GLUT3. 3. Long-term incubation (48 hr), especially with the phenothiazine drugs, was accompanied by a marked reduction in cell growth and proliferation. The rank ordering of the potencies of the drugs was essentially the same for these various effects: fluphenazine > chlorpromazine > clozapine approximately haloperidol. 4. It is suggested that the effects on glucose transport reported here may complicate the interpretation of positron emission tomography (PET) studies that rely on the uptake of radiolabeled glucose analogs to measure the physiological response to these drugs.

Cytotoxicity of conventional and atypical antipsychotic drugs in relation to glucose metabolism

The goal of these studies was to analyze the cytotoxicity of both the conventional and atypical antipsychotic drugs in relation to their effects on glucose metabolism. The drugs were evaluated for their effects on the viability of PC12 cells, which are an established model of neuronal cells in culture. In general, the conventional drugs, such as chlorpromazine, fluphenazine and pimozide, were more toxic than the atypical drugs, including clozapine, quetiapine and risperidone. Olanzapine was unique in that it stimulated cell proliferation in this system. There was a good correlation between the cytotoxicity of a drug and its ability to block glucose transport, although there were some exceptions to this trend. Conventional antipsychotics also affected the expression of glucose transporter proteins in whole cell extracts and at the cell surface. Overall, the data support the notion that many of the antipsychotic drugs associated with the development of movement disorders in patients are cytotoxic for cultured cells.

Model of the 3-D structure of the GLUT3 glucose transporter and molecular dynamics simulation of glucose transport

A molecular model of the three‐dimensional (3‐D) structure of the glucose transport protein, GLUT3, has been derived by homology modeling. The model was built on the basis of structural data from the MscL protein, which is a mechanosensitive ion channel, and general insights from aquaporin (a water permeation pore). Structurally conserved regions were defined by amino acid sequence comparisons, optimum interconnecting loops were selected from the protein databank, and amino (N)‐ and carboxy (C)‐terminal ends of the protein were generated as random coil structures. The model was then subjected to energy minimization and molecular dynamics simulations in the presence of bound substrate (D‐glucose). In the proposed structure of GLUT3, the 12 transmembrane (TM) helices form a right‐hand barrel with a central hydrophilic pore. The pore is shaped like a funnel with dimensions of approximately 5–6 Å by 8 Å at its narrowest point. A network of polar and aromatic amino acids line the pore region and may facilitate the movement of glucose along the channel. A putative binding site for inhibitory ligands, such as forskolin and cytochalasin B, was identified on an intracellular aspect of the protein. Molecular dynamics studies showed that changes in the tilt and flexibility of key TM helices may modulate the opening of the pore to effect glucose transport. The proposed structure of GLUT3 may prove useful in guiding future experiments aimed at more precisely defining various functional regions of the transporter and may encourage efforts to develop models of other complex membrane proteins. Proteins 2001;42:531–541.

Dopamine receptor antagonists modulate glucose uptake in rat pheochromocytoma (PC12) cells

A variety of dopaminergic ligands were evaluated for their ability to alter glucose transport in PC12 cells. Certain antipsychotic drugs which targeted D2 dopamine receptors, such as pimozide, fluphenazine and chlorpromazine, inhibited glucose uptake (with IC50s in the range of 2-40 microM). By contrast, haloperidol and sulpiride (also D2 antagonists) showed marginal activity. The atypical antipsychotic drug, clozapine (a D4 antagonist), also effectively inhibited glucose transport by the cells. Ligands specific for D1 receptors did not interfere with glucose uptake. Time course studies revealed that a short incubation with the drugs (1-5 min) was sufficient to block glucose transport. These findings may have implications for the adverse effects of these drugs and for the interpretation of imaging studies of brain glucose metabolism in patients on antipsychotic medications.

The inhibition of GLUT1 glucose transport and cytochalasin B binding activity by tricyclic antidepressants

Under normal metabolic conditions glucose is an important energy source for the mammalian brain. Positron Emission Tomography studies of the central nervous system have demonstrated that tricyclic antidepressant medications alter cerebral metabolic function. The mode by which these drugs perturb metabolism is unknown. In the present study the interactions of tricyclic antidepressants with the GLUT1 glucose transport protein is examined. Amitriptyline, nortriptyline, desipramine, and imipramine all inhibit the influx of 3-O-methyl glucose into resealed erythrocytes. This inhibition is observed with drug concentrations in the millimolar range. All four antidepressants also noncompetitively displace cytochalasin B binding to GLUT1. The K(I) for this displacement ranges from 0.56 to 1.43 millimolar. This value is in a range greater than that associated with clinical doses and this effect may not be directly applicable to side effects observed with normal use. The observed interaction of these drugs with GLUT1 may reflect an affinity for other glucose-transport or glucose-binding proteins, and may possibly contribute to tricyclic antidepressant toxicity.

Discrimination between mono–and trimethylated cap structures by two isoforms of Caenorhabditis elegans eIF4E

Primitive eukaryotes like Caenorhabditis elegans produce mRNAs capped with either m7GTP or m32,2,7GTP. Caenorhabditis elegans also expresses five isoforms of the cap‐binding protein eIF4E. Some isoforms (e.g. IFE‐3) bind to m7GTP–Sepharose exclusively, whereas others (e.g. IFE‐5) bind to both m7GTP− and m32,2,7GTP–Sepharose. To examine specificity differences, we devised molecular models of the tertiary structures of IFE‐3 and IFE‐5, based on the known structure of mouse eIF4E‐1. We then substituted amino acid sequences of IFE‐5 with homologous sequences from IFE‐3. As few as two changes (N64Y/V65L) converted the cap specificity of IFE‐5 to essentially that of IFE‐3. Molecular dynamics simulations suggested that the width and depth of the cap‐binding cavity were larger in IFE‐5 than in IFE‐3 or the N64Y/V65L variant, supporting a model in which IFE‐3 discriminates against m32,2,7GTP by steric hindrance. Furthermore, the affinity of IFE‐5 (but not IFE‐3) for m32,2,7GTP was reversibly increased when thiol reagents were removed. This was correlated with the formation of a disulfide bond between Cys‐122 and Cys‐126. Thus, translation of m32,2,7GTP‐capped mRNAs may be regulated by intracellular redox state.

Overlap myasthenic syndrome: combined myasthenia gravis and Eaton-Lambert syndrome

A patient with a known history of pernicious anemia had the combined features of autoimmune myasthenia gravis (MG) and the Eaton-Lambert syndrome (ELS). Initially, this patient had all the features typical of MG, and after thymectomy developed all the typical features of ELS. In view of the coexistence of two autoimmune neuromuscular transmission disorders in one patient, we termed this disorder “overlap myasthenic syndrome.”

Antipsychotic drugs activate the C. elegans akt pathway via the DAF-2 insulin/IGF-1 receptor.

The molecular modes of action of antipsychotic drugs are poorly understood beyond their effects at the dopamine D2 receptor. Previous studies have placed Akt signaling downstream of D2 dopamine receptors, and recent data have suggested an association between psychotic illnesses and defective Akt signaling. To characterize the effect of antipsychotic drugs on the Akt pathway, we used the model organism C. elegans, a simple system where the Akt/forkhead box O transcription factor (FOXO) pathway has been well characterized. All major classes of antipsychotic drugs increased signaling through the insulin/Akt/FOXO pathway, whereas four other drugs that are known to affect the central nervous system did not. The antipsychotic drugs inhibited dauer formation, dauer recovery, and shortened lifespan, three biological processes affected by Akt signaling. Genetic analysis showed that AKT-1 and the insulin and insulin-like growth factor receptor, DAF-2, were required for the antipsychotic drugs to increase signaling. Serotonin synthesis was partially involved, whereas the mitogen activated protein kinase (MAPK), SEK-1 is a MAP kinase kinase (MAPKK), and calcineurin were not involved. This is the first example of a common but specific molecular effect produced by all presently known antipsychotic drugs in any biological system. Because untreated schizophrenics have been reported to have low levels of Akt signaling, increased Akt signaling might contribute to the therapeutic actions of antipsychotic drugs.

An ethanol-sensitive variant of the PC12 neuronal cell line: sensitivity to alcohol is associated with increased cell adhesion and decreased glucose accumulation.

A stable variant of the PC12 cell line (PC12.4) has been isolated on the basis of its cell adhesive properties and morphological characteristics. Cells from the PC12.4 subline differ from the parental cell line in that they readily adhere to untreated plastic surfaces and grow individually rather than aggregated in large clusters. When compared to the PC12.1 cell line (original phenotype), PC12.4 cells were found to have a more rapid growth rate (24 h vs. 40 h doubling time) and higher production of lactate but lower glucose metabolism as judged by the accumulation of 3H‐2‐deoxyglucose. Western blot analyses also revealed differences between PC12.1 and PC12.4 cells with respect to the expression of glucose transporters (GLUTs) and the subcellular distribution of the heat shock protein (Hsp) Hsp60. We have reported here that PC12.4 cells were far more sensitive to growth inhibition by ethanol when compared with PC12.1 cells and appeared to be more dependent upon glutamine and serum for cell growth. The cytostatic effects of ethanol were most pronounced when the cells were cultured in medium with low concentrations of serum and glutamine. Thus, there appears to be an interplay between energy metabolism in the cell and the response to ethanol. J Cell Physiol 178:93–101, 1999.