Alzheimer's disease (AD) is a progressive neurodegenerative disorder, which is characterised clinically by dementia and progressive memory loss, and pathologically by neuronal degeneration, plaques (insoluble β-amyloid (Aβ) protein) and neurofibrillary lesions (abnormally phosphorylated tau protein). The mechanisms by which cells die in AD remain largely unknown and controversial. There is some evidence to suggest that cell death in AD brains may occur by apoptosis, and that Aβ might be involved in this process. Apoptosis, a type of cell death characterised by distinct morphological and biochemical features, is often the result of 'programmed cell death' (PCD). Many gene families have been proposed to be involved in the PCD pathway, including the caspase family, inducible transcription factor (ITF) family (including Jun, Fos and Krox genes), and members of the Bcl-2 gene family (including the death promoting gene Bax). It is possible, therefore, that some of these genes may play a role in cell death in AD. The hippocampus is one of the first regions of the brain to be affected in AD, showing cell loss mainly in the CA1-2 pyramidal cell layer. In this thesis, the hippocampus from AD and Control cases has been examined for markers of apoptosis and genes thought to be involved in PCD. In addition, the actions of Aβ, human amylin (a structurally similar protein to Aβ) and the Aβ precursor protein (APP) have been examined in cell culture in an attempt to elucidate their mechanisms of action and relate this to the pathogenesis of AD. AD hippocampi showed increased DNA fragmentation as assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL), but TUNEL-positive cells in AD generally did not exhibit 'typical' apoptotic morphology, and there was no evidence of the oligonucleosomal DNA fragmentation characteristic of apoptosis. This indicates that 'typical' apoptosis may not be the predominant cell death mechanism in AD. However, there was some evidence of atypical 'broken' nuclei, which may represent a form of apoptosis that presents with a different morphology in aging tissue. This study found no conclusive evidence of increased expression of Fos or Jun family members in the CA1 region of AD hippocampi, however there were increased levels of the putative 'apoptosis-specific protein' and krox24 mRNA in this area which could be related to the cell death. There was no change in Bax expression in the CA1 region of AD brains (although increased Bax expression was observed in this region in a rat hypoxic-ischemia model where the CA1 neurons die by apoptosis). However, there was a decrease in Bax expression in the granule cells of AD hippocampi which could be related to the relative preservation of these cells in AD. Bax and ITF expression was observed in tangles, senile plaques and Hirano bodies in AD hippocampi, which may be related to the formation of these features and/or the pathogenesis of AD. There appeared to be changes in the cellular location of proteins in post-mortem tissue that made determination of ITF levels extremely difficult. In addition, patterns of ITF expression differed when different antisera directed at the same protein were used. These observations indicate that caution must be exercised when studying protein changes in post-mortem tissue. Application of insoluble Aβ to cultured cells, and overexpression of APP or familial AD-linked APP mutants in cultured cells, did not cause toxicity or alter c-Jun gene expression. However, human amylin was toxic to cultured cells, and had different effects on c-Jun gene expression depending on the cell type. This shows that structurally similar proteins do not always act by a similar mechanism, and that care must be taken when choosing a cell culture system to study disease-related events. The finding that neither insoluble Aβ nor APP/AD-linked APP mutants caused acute toxicity to cultured cells, coupled with the lack of relationship between TUNEL staining and Aβ deposits in post-mortem AD tissue, indicates that deposited insoluble Aβ and/or increased amounts of Aβ may not represent the toxic event in AD. This thesis provides a detailed investigation of several factors that could be involved in the cell death process in the hippocampus in AD. The results presented find no conclusive evidence for ‘classical’ apoptosis and/or increased ITF expression in the hippocampus in AD, but the changes in expression of krox24 mRNA, ‘apoptosis-specific protein’ and Bax suggest that programmed cell death may well be a mechanism which is involved in the pathogenesis of AD. / Whole document restricted, see Access Instructions file below for details of how to access the print copy. / Related published articles. MacGibbon GA, Cooper GJS, Dragunow M. Acute application of human amylin, unlike β-amyloid peptides, kills undifferentiated PC12 cells by apoptosis. NeuroReport 1997; 8:3945-3950. MacGibbon GA, Lawlor PA, Walton M, et al. Expression of Fos, Jun and Krox family proteins in Alzheimer's disease. Exp Neurol 1997; 147:316-332. MacGibbon GA, Lawlor PA, Sirimanne E et al. Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus. Brain Res 1997; 750:223-234

No description available.

No description available.

No description available.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder, which is characterised clinically by dementia and progressive memory loss, and pathologically by neuronal degeneration, plaques (insoluble β-amyloid (Aβ) protein) and neurofibrillary lesions (abnormally phosphorylated tau protein). The mechanisms by which cells die in AD remain largely unknown and controversial. There is some evidence to suggest that cell death in AD brains may occur by apoptosis, and that Aβ might be involved in this process. Apoptosis, a type of cell death characterised by distinct morphological and biochemical features, is often the result of 'programmed cell death' (PCD). Many gene families have been proposed to be involved in the PCD pathway, including the caspase family, inducible transcription factor (ITF) family (including Jun, Fos and Krox genes), and members of the Bcl-2 gene family (including the death promoting gene Bax). It is possible, therefore, that some of these genes may play a role in cell death in AD. The hippocampus is one of the first regions of the brain to be affected in AD, showing cell loss mainly in the CA1-2 pyramidal cell layer. In this thesis, the hippocampus from AD and Control cases has been examined for markers of apoptosis and genes thought to be involved in PCD. In addition, the actions of Aβ, human amylin (a structurally similar protein to Aβ) and the Aβ precursor protein (APP) have been examined in cell culture in an attempt to elucidate their mechanisms of action and relate this to the pathogenesis of AD. AD hippocampi showed increased DNA fragmentation as assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL), but TUNEL-positive cells in AD generally did not exhibit 'typical' apoptotic morphology, and there was no evidence of the oligonucleosomal DNA fragmentation characteristic of apoptosis. This indicates that 'typical' apoptosis may not be the predominant cell death mechanism in AD. However, there was some evidence of atypical 'broken' nuclei, which may represent a form of apoptosis that presents with a different morphology in aging tissue. This study found no conclusive evidence of increased expression of Fos or Jun family members in the CA1 region of AD hippocampi, however there were increased levels of the putative 'apoptosis-specific protein' and krox24 mRNA in this area which could be related to the cell death. There was no change in Bax expression in the CA1 region of AD brains (although increased Bax expression was observed in this region in a rat hypoxic-ischemia model where the CA1 neurons die by apoptosis). However, there was a decrease in Bax expression in the granule cells of AD hippocampi which could be related to the relative preservation of these cells in AD. Bax and ITF expression was observed in tangles, senile plaques and Hirano bodies in AD hippocampi, which may be related to the formation of these features and/or the pathogenesis of AD. There appeared to be changes in the cellular location of proteins in post-mortem tissue that made determination of ITF levels extremely difficult. In addition, patterns of ITF expression differed when different antisera directed at the same protein were used. These observations indicate that caution must be exercised when studying protein changes in post-mortem tissue. Application of insoluble Aβ to cultured cells, and overexpression of APP or familial AD-linked APP mutants in cultured cells, did not cause toxicity or alter c-Jun gene expression. However, human amylin was toxic to cultured cells, and had different effects on c-Jun gene expression depending on the cell type. This shows that structurally similar proteins do not always act by a similar mechanism, and that care must be taken when choosing a cell culture system to study disease-related events. The finding that neither insoluble Aβ nor APP/AD-linked APP mutants caused acute toxicity to cultured cells, coupled with the lack of relationship between TUNEL staining and Aβ deposits in post-mortem AD tissue, indicates that deposited insoluble Aβ and/or increased amounts of Aβ may not represent the toxic event in AD. This thesis provides a detailed investigation of several factors that could be involved in the cell death process in the hippocampus in AD. The results presented find no conclusive evidence for ‘classical’ apoptosis and/or increased ITF expression in the hippocampus in AD, but the changes in expression of krox24 mRNA, ‘apoptosis-specific protein’ and Bax suggest that programmed cell death may well be a mechanism which is involved in the pathogenesis of AD. / Whole document restricted, see Access Instructions file below for details of how to access the print copy. / Related published articles. MacGibbon GA, Cooper GJS, Dragunow M. Acute application of human amylin, unlike β-amyloid peptides, kills undifferentiated PC12 cells by apoptosis. NeuroReport 1997; 8:3945-3950. MacGibbon GA, Lawlor PA, Walton M, et al. Expression of Fos, Jun and Krox family proteins in Alzheimer's disease. Exp Neurol 1997; 147:316-332. MacGibbon GA, Lawlor PA, Sirimanne E et al. Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus. Brain Res 1997; 750:223-234

Alzheimer's disease (AD) is a progressive neurodegenerative disorder, which is characterised clinically by dementia and progressive memory loss, and pathologically by neuronal degeneration, plaques (insoluble β-amyloid (Aβ) protein) and neurofibrillary lesions (abnormally phosphorylated tau protein). The mechanisms by which cells die in AD remain largely unknown and controversial. There is some evidence to suggest that cell death in AD brains may occur by apoptosis, and that Aβ might be involved in this process. Apoptosis, a type of cell death characterised by distinct morphological and biochemical features, is often the result of 'programmed cell death' (PCD). Many gene families have been proposed to be involved in the PCD pathway, including the caspase family, inducible transcription factor (ITF) family (including Jun, Fos and Krox genes), and members of the Bcl-2 gene family (including the death promoting gene Bax). It is possible, therefore, that some of these genes may play a role in cell death in AD. The hippocampus is one of the first regions of the brain to be affected in AD, showing cell loss mainly in the CA1-2 pyramidal cell layer. In this thesis, the hippocampus from AD and Control cases has been examined for markers of apoptosis and genes thought to be involved in PCD. In addition, the actions of Aβ, human amylin (a structurally similar protein to Aβ) and the Aβ precursor protein (APP) have been examined in cell culture in an attempt to elucidate their mechanisms of action and relate this to the pathogenesis of AD. AD hippocampi showed increased DNA fragmentation as assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL), but TUNEL-positive cells in AD generally did not exhibit 'typical' apoptotic morphology, and there was no evidence of the oligonucleosomal DNA fragmentation characteristic of apoptosis. This indicates that 'typical' apoptosis may not be the predominant cell death mechanism in AD. However, there was some evidence of atypical 'broken' nuclei, which may represent a form of apoptosis that presents with a different morphology in aging tissue. This study found no conclusive evidence of increased expression of Fos or Jun family members in the CA1 region of AD hippocampi, however there were increased levels of the putative 'apoptosis-specific protein' and krox24 mRNA in this area which could be related to the cell death. There was no change in Bax expression in the CA1 region of AD brains (although increased Bax expression was observed in this region in a rat hypoxic-ischemia model where the CA1 neurons die by apoptosis). However, there was a decrease in Bax expression in the granule cells of AD hippocampi which could be related to the relative preservation of these cells in AD. Bax and ITF expression was observed in tangles, senile plaques and Hirano bodies in AD hippocampi, which may be related to the formation of these features and/or the pathogenesis of AD. There appeared to be changes in the cellular location of proteins in post-mortem tissue that made determination of ITF levels extremely difficult. In addition, patterns of ITF expression differed when different antisera directed at the same protein were used. These observations indicate that caution must be exercised when studying protein changes in post-mortem tissue. Application of insoluble Aβ to cultured cells, and overexpression of APP or familial AD-linked APP mutants in cultured cells, did not cause toxicity or alter c-Jun gene expression. However, human amylin was toxic to cultured cells, and had different effects on c-Jun gene expression depending on the cell type. This shows that structurally similar proteins do not always act by a similar mechanism, and that care must be taken when choosing a cell culture system to study disease-related events. The finding that neither insoluble Aβ nor APP/AD-linked APP mutants caused acute toxicity to cultured cells, coupled with the lack of relationship between TUNEL staining and Aβ deposits in post-mortem AD tissue, indicates that deposited insoluble Aβ and/or increased amounts of Aβ may not represent the toxic event in AD. This thesis provides a detailed investigation of several factors that could be involved in the cell death process in the hippocampus in AD. The results presented find no conclusive evidence for ‘classical’ apoptosis and/or increased ITF expression in the hippocampus in AD, but the changes in expression of krox24 mRNA, ‘apoptosis-specific protein’ and Bax suggest that programmed cell death may well be a mechanism which is involved in the pathogenesis of AD. / Whole document restricted, see Access Instructions file below for details of how to access the print copy. / Related published articles. MacGibbon GA, Cooper GJS, Dragunow M. Acute application of human amylin, unlike β-amyloid peptides, kills undifferentiated PC12 cells by apoptosis. NeuroReport 1997; 8:3945-3950. MacGibbon GA, Lawlor PA, Walton M, et al. Expression of Fos, Jun and Krox family proteins in Alzheimer's disease. Exp Neurol 1997; 147:316-332. MacGibbon GA, Lawlor PA, Sirimanne E et al. Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus. Brain Res 1997; 750:223-234

Alzheimer's disease (AD) is a progressive neurodegenerative disorder, which is characterised clinically by dementia and progressive memory loss, and pathologically by neuronal degeneration, plaques (insoluble β-amyloid (Aβ) protein) and neurofibrillary lesions (abnormally phosphorylated tau protein). The mechanisms by which cells die in AD remain largely unknown and controversial. There is some evidence to suggest that cell death in AD brains may occur by apoptosis, and that Aβ might be involved in this process. Apoptosis, a type of cell death characterised by distinct morphological and biochemical features, is often the result of 'programmed cell death' (PCD). Many gene families have been proposed to be involved in the PCD pathway, including the caspase family, inducible transcription factor (ITF) family (including Jun, Fos and Krox genes), and members of the Bcl-2 gene family (including the death promoting gene Bax). It is possible, therefore, that some of these genes may play a role in cell death in AD. The hippocampus is one of the first regions of the brain to be affected in AD, showing cell loss mainly in the CA1-2 pyramidal cell layer. In this thesis, the hippocampus from AD and Control cases has been examined for markers of apoptosis and genes thought to be involved in PCD. In addition, the actions of Aβ, human amylin (a structurally similar protein to Aβ) and the Aβ precursor protein (APP) have been examined in cell culture in an attempt to elucidate their mechanisms of action and relate this to the pathogenesis of AD. AD hippocampi showed increased DNA fragmentation as assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL), but TUNEL-positive cells in AD generally did not exhibit 'typical' apoptotic morphology, and there was no evidence of the oligonucleosomal DNA fragmentation characteristic of apoptosis. This indicates that 'typical' apoptosis may not be the predominant cell death mechanism in AD. However, there was some evidence of atypical 'broken' nuclei, which may represent a form of apoptosis that presents with a different morphology in aging tissue. This study found no conclusive evidence of increased expression of Fos or Jun family members in the CA1 region of AD hippocampi, however there were increased levels of the putative 'apoptosis-specific protein' and krox24 mRNA in this area which could be related to the cell death. There was no change in Bax expression in the CA1 region of AD brains (although increased Bax expression was observed in this region in a rat hypoxic-ischemia model where the CA1 neurons die by apoptosis). However, there was a decrease in Bax expression in the granule cells of AD hippocampi which could be related to the relative preservation of these cells in AD. Bax and ITF expression was observed in tangles, senile plaques and Hirano bodies in AD hippocampi, which may be related to the formation of these features and/or the pathogenesis of AD. There appeared to be changes in the cellular location of proteins in post-mortem tissue that made determination of ITF levels extremely difficult. In addition, patterns of ITF expression differed when different antisera directed at the same protein were used. These observations indicate that caution must be exercised when studying protein changes in post-mortem tissue. Application of insoluble Aβ to cultured cells, and overexpression of APP or familial AD-linked APP mutants in cultured cells, did not cause toxicity or alter c-Jun gene expression. However, human amylin was toxic to cultured cells, and had different effects on c-Jun gene expression depending on the cell type. This shows that structurally similar proteins do not always act by a similar mechanism, and that care must be taken when choosing a cell culture system to study disease-related events. The finding that neither insoluble Aβ nor APP/AD-linked APP mutants caused acute toxicity to cultured cells, coupled with the lack of relationship between TUNEL staining and Aβ deposits in post-mortem AD tissue, indicates that deposited insoluble Aβ and/or increased amounts of Aβ may not represent the toxic event in AD. This thesis provides a detailed investigation of several factors that could be involved in the cell death process in the hippocampus in AD. The results presented find no conclusive evidence for ‘classical’ apoptosis and/or increased ITF expression in the hippocampus in AD, but the changes in expression of krox24 mRNA, ‘apoptosis-specific protein’ and Bax suggest that programmed cell death may well be a mechanism which is involved in the pathogenesis of AD. / Whole document restricted, see Access Instructions file below for details of how to access the print copy. / Related published articles. MacGibbon GA, Cooper GJS, Dragunow M. Acute application of human amylin, unlike β-amyloid peptides, kills undifferentiated PC12 cells by apoptosis. NeuroReport 1997; 8:3945-3950. MacGibbon GA, Lawlor PA, Walton M, et al. Expression of Fos, Jun and Krox family proteins in Alzheimer's disease. Exp Neurol 1997; 147:316-332. MacGibbon GA, Lawlor PA, Sirimanne E et al. Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus. Brain Res 1997; 750:223-234

No description available.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder, which is characterised clinically by dementia and progressive memory loss, and pathologically by neuronal degeneration, plaques (insoluble β-amyloid (Aβ) protein) and neurofibrillary lesions (abnormally phosphorylated tau protein). The mechanisms by which cells die in AD remain largely unknown and controversial. There is some evidence to suggest that cell death in AD brains may occur by apoptosis, and that Aβ might be involved in this process. Apoptosis, a type of cell death characterised by distinct morphological and biochemical features, is often the result of 'programmed cell death' (PCD). Many gene families have been proposed to be involved in the PCD pathway, including the caspase family, inducible transcription factor (ITF) family (including Jun, Fos and Krox genes), and members of the Bcl-2 gene family (including the death promoting gene Bax). It is possible, therefore, that some of these genes may play a role in cell death in AD. The hippocampus is one of the first regions of the brain to be affected in AD, showing cell loss mainly in the CA1-2 pyramidal cell layer. In this thesis, the hippocampus from AD and Control cases has been examined for markers of apoptosis and genes thought to be involved in PCD. In addition, the actions of Aβ, human amylin (a structurally similar protein to Aβ) and the Aβ precursor protein (APP) have been examined in cell culture in an attempt to elucidate their mechanisms of action and relate this to the pathogenesis of AD. AD hippocampi showed increased DNA fragmentation as assessed by TdT-mediated dUTP-biotin nick end labelling (TUNEL), but TUNEL-positive cells in AD generally did not exhibit 'typical' apoptotic morphology, and there was no evidence of the oligonucleosomal DNA fragmentation characteristic of apoptosis. This indicates that 'typical' apoptosis may not be the predominant cell death mechanism in AD. However, there was some evidence of atypical 'broken' nuclei, which may represent a form of apoptosis that presents with a different morphology in aging tissue. This study found no conclusive evidence of increased expression of Fos or Jun family members in the CA1 region of AD hippocampi, however there were increased levels of the putative 'apoptosis-specific protein' and krox24 mRNA in this area which could be related to the cell death. There was no change in Bax expression in the CA1 region of AD brains (although increased Bax expression was observed in this region in a rat hypoxic-ischemia model where the CA1 neurons die by apoptosis). However, there was a decrease in Bax expression in the granule cells of AD hippocampi which could be related to the relative preservation of these cells in AD. Bax and ITF expression was observed in tangles, senile plaques and Hirano bodies in AD hippocampi, which may be related to the formation of these features and/or the pathogenesis of AD. There appeared to be changes in the cellular location of proteins in post-mortem tissue that made determination of ITF levels extremely difficult. In addition, patterns of ITF expression differed when different antisera directed at the same protein were used. These observations indicate that caution must be exercised when studying protein changes in post-mortem tissue. Application of insoluble Aβ to cultured cells, and overexpression of APP or familial AD-linked APP mutants in cultured cells, did not cause toxicity or alter c-Jun gene expression. However, human amylin was toxic to cultured cells, and had different effects on c-Jun gene expression depending on the cell type. This shows that structurally similar proteins do not always act by a similar mechanism, and that care must be taken when choosing a cell culture system to study disease-related events. The finding that neither insoluble Aβ nor APP/AD-linked APP mutants caused acute toxicity to cultured cells, coupled with the lack of relationship between TUNEL staining and Aβ deposits in post-mortem AD tissue, indicates that deposited insoluble Aβ and/or increased amounts of Aβ may not represent the toxic event in AD. This thesis provides a detailed investigation of several factors that could be involved in the cell death process in the hippocampus in AD. The results presented find no conclusive evidence for ‘classical’ apoptosis and/or increased ITF expression in the hippocampus in AD, but the changes in expression of krox24 mRNA, ‘apoptosis-specific protein’ and Bax suggest that programmed cell death may well be a mechanism which is involved in the pathogenesis of AD. / Whole document restricted, see Access Instructions file below for details of how to access the print copy. / Related published articles. MacGibbon GA, Cooper GJS, Dragunow M. Acute application of human amylin, unlike β-amyloid peptides, kills undifferentiated PC12 cells by apoptosis. NeuroReport 1997; 8:3945-3950. MacGibbon GA, Lawlor PA, Walton M, et al. Expression of Fos, Jun and Krox family proteins in Alzheimer's disease. Exp Neurol 1997; 147:316-332. MacGibbon GA, Lawlor PA, Sirimanne E et al. Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus. Brain Res 1997; 750:223-234

<div><b>Background:</b> Bupropion, an atypical antidepressant and smoking cessation aid, is associated with wide inter-subject variability in its efficacy and safety. Variability in response to bupropion therapy is thought to be driven by variability in metabolism. Bupropion undergoes complex phase 1 and 2 stereoselective metabolism. Though bupropion`s pharmacology is not fully understood, much of it is thought to be due to its metabolites, specially, S, S-hydroxybupropion. In vitro studies (functional assays measuring IC50 at dopamine transporter-DAT, norepinephrine transporter-NET, various subtypes of nicotinic receptors-nAChR) and mouse models (forced swim test to assess antidepressant effect, antinociceptive models to assess antagonism of nicotine effects) indicate S, S-hydroxybupropion to contribute more towards efficacy as an antidepressant and smoking cessation aid than racemic bupropion and R, R-hydroxybupropion, respectively. Both pharmacokinetics (PK) and pharmacodynamics (PD) of bupropion and its metabolites are complex and reported to be stereoselective. As bupropion is known to act on multiple central nervous system (CNS) targets (DAT, NET nAChR), understanding CNS disposition (target site) is critical to explain variability in bupropion`s therapeutic and toxic effects. </div><div><b>Objective: </b>The objective of our study was to characterize the exposure of bupropion enantiomers and corresponding phase 1 metabolite diastereomers in plasma and brain in a surrogate non-clinical species, and to subsequently develop animal-to-human-translational population-PK and Physiologically Based PK (PBPK) models to predict human brain concentrations of bupropion and its active metabolite S, S-hydroxybupropion. Application of these PK modeling approaches to map the time course of unbound brain concentration can then be compared to in vitro potency measures at DAT, NET and nAChRs to predict target engagement over time (PD). Establishing relationships between plasma PK, target site PK along with PD would elucidate possible cause(s) of inter-patient variability to bupropion therapy. </div><div><b>Methods: </b>The first step towards development of a CNS model was to identify a nonclinical species with phase 1 metabolism closest to humans. To accomplish this, hepatic microsomal incubations of four species-rat, mouse, non-human primates (NHPs) and humans were conducted separately for the R- and S-bupropion enantiomers, and the formation of enantiomer-specific metabolites was determined using LC-MS/MS. Intrinsic formation clearance (CLint) of metabolites across the four species (rats, mice, NHPs, humans) was determined from the formation rate versus substrate concentration relationship. </div><div>Racemic bupropion (10 mg/kg) and preformed S, S-hydroxybupropion (2 mg/kg) were administered subcutaneously to adult male Sprague Dawley rats (n = 24/compound). Brain and plasma were collected from rats (n = 3) at eight time points for 6 hours and analyzed using a chiral LC-MS/MS method. Rat plasma protein and brain homogenate binding studies were conducted for all analytes to correct for unbound fraction using equilibrium dialysis method.</div><div>A plasma-brain compartmental pharmacokinetic approach was used to describe the blood–brain-barrier transport of both bupropion and S, S-hydroxybupropion. Also, a 2-compartment permeability-limited brain model consisting of brain blood, brain mass compartments was developed and incorporated into a whole body physiologically-based pharmacokinetic (PBPK) parent-metabolite model for bupropion and S, S-hydroxybupropion. Both population PK and PBPK modeling approaches were subsequently translated to humans to predict human plasma and brain site exposure and its relationship to DAT and NET IC50 potencies.</div><div><b>Results: </b>The total clearance of S-bupropion was higher than that of R-bupropion in monkey and human liver microsomes. The contribution of hydroxybupropion to the total racemic bupropion clearance was 38%, 62%, 17%, and 96% in human, monkey, rat, and mouse, respectively. In the same species order, threohydrobupropion contributed 53%, 23%, 17%, and 3%, and erythrohydrobupropion contributed 9%, 14%, 66%, and 1.3%, respectively, to racemic bupropion clearance. Hepatic microsomal incubation studies indicated non-human primates to be the appropriate species to model CNS disposition. However, the cost and limited pharmacokinetic and pharmacodynamic data in NHPs were insurmountable barriers to conducting in vivo studies in NHPs. After considering multiple factors, such as the formation of reductive metabolites (higher in rats than mice), which are also thought to contribute to bupropion`s therapeutic efficacy, availability of microdialysis data measuring bupropion and dopamine, norepinephrine levels in brain extracellular fluid (ECF) and other in vitro potency evaluations in rats, rat was chosen as the surrogate species to model bupropion`s disposition.</div><div>In rats, unbound plasma and brain exposures and plasma clearances of both R and S-bupropion were similar. The exposure to parent was higher (50 to 100-fold) than to metabolites. The exposure of oxidative metabolites (R, R- and S, S-hydroxybupropion) was 2 to 3-fold higher in brain and plasma than reductive metabolites (R, R- and S, S-threohydrobupropion, S, R- and R, S-erythrohydrobupropion). Hepatic clearances of R- and S-bupropion scaled from in vitro rat hepatic microsomal incubation studies were 3-fold and 25-fold lower than their respective in vivo unbound apparent clearances. This could possibly be due to substantial contribution of metabolic pathways not characterized in this in vivo study and/or possible extrahepatic disposition in the rat. The unbound brain to unbound plasma AUC0-6h ratio (Kp,uu) of R- and S-bupropion were 0.43 and 0.38 respectively. Kp,uu of oxidative metabolites (R, R- and S, S-hydroxybupropion) and reductive metabolites (R, R- and S, S-threohydrobupropion) were close to 1. Kp,uu of S, R-erythrohydrobupropion was 0.43 and that of pre-formed S, S-hydroxybupropion was 5.</div><div>With respect to population PK modeling of both bupropion and S, S-hydroxybupropion, a plasma-brain compartmental model structure with time dependent change in brain influx clearance was required to adequately characterize the BBB transport of parent and this active metabolite. Using a physiologically-based pharmacokinetic model (PBPK) approach too, incorporation of active efflux and carrier mediated uptake terms in addition to passive permeability was necessary to adequately characterize brain disposition of bupropion and S, S-hydroxybupropion. Both modeling approaches (population-PK and PBPK) when translated to humans indicated that the predicted human brain exposures fall below the reported DAT and NET IC50 measures of bupropion and S, S-hydroxybupropion. </div><div><b>Conclusion: </b>Specific to our work in the rat, the discrepancy between in vitro scaled hepatic clearance and in vivo plasma clearance of R and S-bupropion suggests alternative non-CYP mediated clearance pathways and/or extra hepatic disposition of bupropion. Both translational PK models indicate active process such as efflux transporter or carrier mediated uptake could be involved in bupropion`s disposition in the brain. Variability in expression of these speculated active/carrier mediated transporters could possibly cause variability in response. Also, other CNS targets could contribute to bupropion`s therapeutic efficacy, elucidation of which would require further investigation.</div><div><br></div>