The experimental Alzheimer drug phenserine: preclinical pharmacokinetics and pharmacodynamics.

Phenserine, a phenylcarbamate of physostigmine, is a new potent and highly selective acetylcholinesterase (AChE) inhibitor, with a > 50-fold activity versus butyrylcholinesterase (BChE), in clinical trials for the treatment of Alzheimer's disease (AD). Compared to physostigmine and tacrine, it is less toxic and robustly enhances cognition in animal models. To determine the time-dependent effects of phenserine on cholinergic function, AChE activity, brain and plasma drug levels and brain extracellular acetylcholine (ACh) concentrations were measured in rats before and after phenserine administration. Additionally, its maximum tolerated dose, compared to physostigmine and tacrine, was determined. Following i.v. dosing, brain drug levels were 10-fold higher than those achieved in plasma, peaked within 5 min and rapidly declined with half-lives of 8.5 and 12.6 min, respectively. In contrast, a high (> 70%) and long-lasting inhibition of AChE was achieved (half-life > 8.25 h). A comparison between the time-dependent plasma AChE inhibition achieved after similar oral and i.v. doses provided an estimate of oral bioavailability of 100%. Striatal, in vivo microdialysis in conscious, freely-moving phenserine-treated rats demonstrated > 3-fold rise in brain ACh levels. Phenserine thus is rapidly absorbed and cleared from the body, but produces a long-lasting stimulation of brain cholinergic function at well tolerated doses and hence has superior properties as a drug candidate for AD. It selectively inhibits AChE, minimizing potential BChE side effects. Its long duration of action, coupled with its short pharmacokinetic half-life, reduces dosing frequency, decreases body drug exposure and minimizes the dependence of drug action on the individual variations of drug metabolism commonly found in the elderly.

Neuroendocrine responses to intravenous infusion of physostigmine in patients with Alzheimer disease.

We have reported that physostigmine, a reversible cholinesterase inhibitor, enhances verbal memory in patients with Alzheimer disease (AD). To elucidate the mechanism of cognition enhancement, plasma hormones were measured during high-dose acute and low-dose chronic steady-state intravenous infusions of physostigmine in nine subjects with AD. High-dose hormone responses were measured during and for 24 h after the infusion of physostigmine 1-1.5 mg over 45-60 min. Chronic responses were measured during continuous intravenous infusions of physostigmine at doses (0.5-25 mg/day) that escalated over 2 weeks, and then during 1 week infusion of the dose that optimized cognition (2-12 mg/day) or placebo administered in a randomized, double-blind, cross-over design. A replicable improvement in verbal memory was found in five subjects. High-dose physostigmine infusion that produced noxious side effects resulted in significant elevation above baseline in plasma levels of adrenocorticotrophic hormone (ACTH) (p = 0.0001), cortisol (p = 0.0001), and beta-endorphin (p = 0.0001). Chronic physostigmine administration, in the absence of adverse effects, produced no significant elevation in ACTH (p = 0.08), cortisol (p = 0.70), or beta-endorphin (p = 0.82). These results indicate that high-dose physostigmine activates the hypothalamic-pituitary-adrenal (HPA) axis, likely representing a "stress response." In contrast, cognition-enhancing doses do not produce a peripheral corticosteroid response. Thus, physostigmine-induced memory improvement is independent of the activation of the HPA axis.

Clinical pharmacokinetics of arecoline in subjects with Alzheimer's disease.

OBJECTIVE: To study the pharmacokinetics and pharmacodynamics of intravenously administered arecoline in subjects with Alzheimer's disease. METHODS: Plasma arecoline concentrations were measured during and after high-dose (i.e., 5 mg intravenously over 30 minutes) and up to 2 weeks of continuous multiple-dose steady-state intravenous infusions of arecoline in 15 subjects with mild to moderate Alzheimer's disease. During multiple-dose infusions, the dose of arecoline was escalated from 0.5 to 40 mg/day. Psychometric tests were administered at baseline and every other dose to determine an "optimal dose" for each subject. This dose then was administered for 1 week using a randomized, placebo-controlled, double blind, crossover design. Plasma drug concentrations were measured by GC-MS. RESULTS: The optimal dose of arecoline varied fourfold across subjects (4 mg/day, n = 6; 16 mg/day, n = 3) with mean plasma half-lives of 0.95 +/- 0.54 and 9.3 +/- 4.5 (SD) minutes. Clearance and volume of distribution were 13.6 +/- 5.8 L/min and 205 +/- 170 (SD) L, respectively. At the dose that optimized memory, the mean plasma level was 0.31 +/- 0.14 (SD) ng/ml, and it predicted the optimal dose in all subjects. CONCLUSIONS: Because optimal dose variation is due to differing plasma kinetics, the plasma arecoline level measured at a single infusion rate can be used to choose the optimal dose for memory enhancement in patients with Alzheimer's disease.

Differential response to the cholinergic agonist arecoline among different cognitive modalities in Alzheimer's disease.

Nine patients with possible or probable dementia of the Alzheimer type were tested on nine cognitive tests prior to (two times) and during continuous intravenous administration of five different doses of the muscarinic cholinergic agonist arecoline (1, 4, 16, 28, and 40 mg/day). The present analysis examined whether improvement on cognitive testing for each patient during arecoline treatment was most likely to occur at the same dose for all tests or whether different test scores improved at different doses of arecoline. Results indicated there were significant differences among tests in the dose at which most patients showed improved cognitive performance. These differences may have therapeutic significance, as verbal ability tended to improve at low doses of arecoline, whereas attention and visuospatial ability tended to improve at higher doses of arecoline.

Clinical pharmacokinetics of physostigmine in patients with Alzheimer's disease.

OBJECTIVE: To study the pharmacokinetic and pharmacodynamic properties of physostigmine in subjects with Alzheimer's disease. METHODS: Plasma physostigmine concentration and butyrylcholinesterase inhibition were measured in blood samples collected during and after a single high-dose (1 to 1.5 mg for 45 to 60 minutes) and a sustained low-dose steady-state intravenous infusion in nine subjects with Alzheimer's disease. Escalating doses (0.5 to 25 mg/day) were administered during a 2-week period. A dose (2 to 12 mg/day) that optimized cognition in each subject was identified and then administered in a randomized, double-blind, placebo-controlled crossover design for 1 week. RESULTS: The elimination half-life of physostigmine was 16.4 +/- 3.2 (SE) minutes. Clearance and volume of distribution were 7.7 +/- 0.9 (SE) L/min and 2.4 +/- 0.6 (SE) L/kg, respectively. Butyrylcholinesterase inhibition half-life was 83.7 +/- 5.2 (SE) minutes. During sustained steady-state infusion, plasma physostigmine concentration (r = 0.95) and butyrylcholinesterase inhibition (r = 0.99) were linearly correlated with the dose. In five cognitive responders, the memory enhancement was significantly correlated (r = 0.86; p < 0.05) with butyrylcholinesterase inhibition. CONCLUSIONS: These results showed that, in cognitive responders, memory enhancement by physostigmine in Alzheimer's disease is correlated directly to the magnitude of plasma cholinesterase inhibition. Furthermore, during single-dose conditions, the dynamic half-life is five-fold longer than the kinetic half-life.

Treatment of Alzheimer disease by continuous intravenous infusion of physostigmine.

Physostigmine, a reversible and nonselective cholinesterase inhibitor, administered by steady-state, continuous intravenous infusion to carefully selected subjects with mild-moderate Alzheimer disease, produced significant but modest improvement in memory in five of nine subjects. Drug dosing was limited by the occurrence of adverse effects. Apparent tolerance to adverse effects was observed in two subjects when the dose of physostigmine was escalated slowly over at least 2 weeks. Steady-state cholinesterase inhibition by physostigmine appears to produce sustained cognitive improvement in some subjects with Alzheimer disease without substantially altering its therapeutic index.

Neuroendocrine responses to intravenous infusion of arecoline in patients with Alzheimer's disease.

We have reported that arecoline, a muscarinic receptor agonist replicably enhanced verbal memory in five of nine subjects with Alzheimer's disease (AD). To investigate the mechanism of cognitive improvement, circulating hormone measurements were made during high-dose acute and low-dose chronic intravenous (i.v.) arecoline administration to AD patients. Acute hormone responses were measured during, and for 6 h after, infusion of arecoline 5 mg i.v. over 30 min. Chronic responses were measured in cognitive responders during continuous i.v. infusion of arecoline escalating over 2 weeks (0.5-40 mg/day) and then during a 1 week infusion of the dose optimizing cognition (4-16 mg/day). Acute arecoline administered to 14 subjects produced unpleasant side-effects (e.g. nausea, vomiting), mean adrenocorticotrophic hormone (p = .0006), cortisol (p = .0001) and beta-endorphin (p = .0001) levels were elevated. During chronic arecoline treatment, no side-effects occurred and plasma cortisol, adrenocorticotrophic hormone and beta-endorphin levels were unchanged in nine subjects overall and in five cognitive responders. Thus, high-dose arecoline activates the hypothalamic-pituitary-adrenal (HPA) axis and may increase other anterior pituitary hormone levels, likely representing a 'stress response', but cognition-enhancing, low doses of arecoline do not produce a glucocorticoid response. Hence, arecoline-induced memory improvement is not due to the induction of 'stress' nor to the elevation of peripheral corticosteroid levels.

Effect of acute and chronic arecoline treatment on cerebral metabolism and blood flow in the conscious rat.

Treatment with the muscarinic agonist arecoline improves memory retention in patients with Alzheimer's disease (AD). In animal models, arecoline selectively increases local cerebral glucose utilization (LCGU). We examined (1) whether these focal increases in metabolism were coupled to local cerebral blood flow (LCBF) and (2) whether the effect of arecoline on LCGU and LCBF was dependent upon duration of drug administration. In groups of young Fischer-344 rats, LCGU and LCBF were determined in 59 brain regions by the [14C]2-deoxyglucose and the [14C]iodoantipyrine autoradiographic methods following either the acute administration of arecoline (2 mg/kg and 15 mg/kg) or the chronic three week administration of arecoline (50 mg/kg/day). In general, LCBF correlated closely with LCGU following arecoline 2 mg/kg administration, but heterogeneous regions were present. Following treatment with arecoline 15 mg/kg, the two parameters became uncoupled with LCBF increasing disproportionately in relation to LCGU. Coupling between LCBF and LCGU was preserved during chronic arecoline treatment (50 mg/kg/day) but some regions, such as the hippocampus, were uncoupled with LCGU increasing to a greater extent than LCBF. Thus, we demonstrate that acute and chronic administration of arecoline can differentially modulate LCBF and LCGU. Since clinical administration of arecoline can improve cognitive function in patients with AD, understanding the ability of arecoline to selectively alter LCBF and LCGU in regions such as the hippocampus may offer insight into the pathophysiology of AD and provide direction for the development of definitive therapy for neurodegenerative disorders.

The monosialoganglioside GM1 dose-dependently reduces regional cerebral metabolic rates for glucose in awake rats.

Using the quantitative autoradiographic [14C]2-deoxyglucose technique, regional cerebral metabolic rates for glucose (rCMRglc) were measured in awake male Fischer-344 rats at 1, 2, 3, 4 and 6 h after administration of GM1 30 mg/kg and at 3 h after GM1 150 or 300 mg/kg. GM1 is a natural compound that is able to prevent neuron degeneration induced by exposure to excitatory amino acids in vitro and by ischemia or neurotoxins in vivo. GM1 30 mg/kg, a dose very effective in preventing excitatory amino acid-induced neurotoxicity, produced minimal rCMRglc change over a 6 h period. GM1 150 and 300 mg/kg reduced rCMRglc, in 14 (31%) and in 29 (64%) brain regions, respectively. Maximal metabolic effects occurred in hippocampal areas which possess, in specific subfields, the highest brain concentrations of different excitatory amino acid receptor subtypes. This finding suggests an effect by GM1 on postreceptor mechanisms common to different excitatory amino acids.

Sustained cortical metabolic responsivity to physostigmine after nucleus basalis magnocellularis ablation in rats.

Unilateral nucleus basalis magnocellularis (NBM) ablation, which causes partial cholinergic denervation of the ipsilateral anterior neocortex, results in an acute but transient depression of regional cerebral metabolic rates for glucose (rCMRglc) in deafferented areas; rCMRglc normalizes within 2 weeks. To seek possible compensatory changes in cholinergic mechanisms following NBM ablation that could lead to rapid metabolic normalization, we studied rCMRglc responses to the receptor agonists nicotine and arecoline and the cholinesterase inhibitor physostigmine in rats at 2 weeks after unilateral NBM destruction. Physostigmine increased rCMRglc in 10 of 30 cortical areas contralateral to the NBM lesion. Compared to the unlesioned side, rCMRglc after physostigmine in the lesioned cortex was significantly lower in 2, significantly higher in 1 and not different (P < 0.05) in 27 areas. Neither arecoline nor nicotine treatment produced rCMRglc asymmetry in lesioned rats. These results demonstrate that responsivity to physostigmine is maintained in most regions of the rat neocortex after extrinsic cholinergic denervation by NBM ablation. This adaptive response appears not to result from cholinergic receptor upregulation and may reflect instead reorganization of cholinergic synapses.

Memory improvement without toxicity during chronic, low dose intravenous arecoline in Alzheimer's disease.

Arecoline, a cholinergic agonist, administered at low doses by continuous intravenous infusion for up to 2 weeks, significantly and replicably improved memory in five of nine subjects with mild-moderate Alzheimer's disease. During dose finding, performance on a verbal memory task improved with an inverted U-shaped relation to dose. Six of nine subjects were classified as responders. During blinded, placebo-controlled, individualized optimal dosing for 5 days, verbal memory again improved in five of six responders but not in any non-responder. No adverse drug effects occurred. Arecoline, and possibly other cholinergic agonists, can safely improve memory in Alzheimer's disease at doses much lower than previously studied.

Time- and dose-dependent effects of the serotonergic agent quipazine on regional cerebral metabolism in rats.

The time course and the relation to dose of regional cerebral metabolic rates for glucose (rCMRglc) were measured in awake male adult Fischer-344 rats after administration of quipazine, a serotonin 5-HT2-3 receptor agonist. rCMRglc was determined, using the quantitative autoradiographic [14C]deoxyglucose technique, in 92 brain regions at 30, 60, 90 and 120 min after quipazine 20 mg/kg i.p. and at 60 min after quipazine 5 mg/kg i.p. Peak metabolic effects were observed 60 min after quipazine 20 mg/kg i.p. when rCMRglc was significantly elevated in 27 (29%) brain regions (mean rise 17%). Quipazine increased rCMRglc in brain regions with high densities of 5-HT3 receptors (area postrema, olfactory tubercle, amygdala), in dopaminergic nuclei (substantia nigra pars compacta and pars reticulata) and terminal fields of their projections (zona incerta, subthalamic nucleus, preoptic magnocellular area, nucleus of facial nerve). The topographic distribution and direction of rCMRglc changes induced by quipazine are different from those produced by the 5-HT2 agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane and, consistent with the pharmacological and binding properties of quipazine, suggest a preferential activation of 5-HT3 receptors.

Cerebral metabolic responses to meta-chlorophenylpiperazine are reduced during its chronic administration to young and aged rats.

The effects of the 5-HT agonist meta-chlorophenylpiperazine (MCPP) on regional cerebral metabolic rates for glucose (rCMRglc) were measured in 3- and 24-month-old rats that were not pretreated or were pretreated for 2 weeks with continuous infusion of saline or MCPP. rCMRglc were measured using the quantitative autoradiographic [14C]2-deoxy-D-glucose technique in 71 brain regions at 15 min after acute administration of MCPP 2.5 mg/kg. In the absence of chronic pretreatment, intraperitoneal MCPP 2.5 mg/kg produced widespread rCMRglc reductions (41 brain areas) in 3-month-old rats and more limited rCMRglc decreases (8 brain areas) in 24-month-old rats. After chronic treatment, MCPP failed to reduce rCMRglc in any region of either group of rats. These findings indicate that mechanisms of downregulation of response to MCPP are functional in young and aged rats and suggest that the age-related reduction in rCMRglc responses to acute MCPP in non-pretreated animals may be due to compensation for age-related losses of 5-HT terminals.

Age-dependent cerebral metabolic effects of unilateral nucleus basalis magnocellularis ablation in rats.

To investigate the age-dependent functional importance of cholinergic neocortical inputs, and to explore whether cortical cholinergic denervation in aged animals might better model the cerebral metabolic changes of Alzheimer's disease, the effects of unilateral ablation of the nucleus basalis magnocellularis (NBM) on cerebral glucose metabolism were studied in young and aged rats. Regional cerebral metabolic rates for glucose (rCMRglc) were determined, using the [14C]deoxyglucose method, in 48 brain regions of 3- and 24-month old Fischer-344 rats at 3, 7, 14 and 28 days after stereotaxic injection of ibotenate into the right NBM, and in sham-operated animals at 3 and 14 days later. For both ages the peak effect of unilateral NBM ablation occurred 3 days later: in young rats, rCMRglc was significantly reduced (compared to the contralateral side) in all 24 anterior cortical areas examined (mean decline 20%), whereas in aged animals, only 9 of 24 areas showed a significant decline in glucose utilization, and the magnitude of rCMRglc reduction (9%) was smaller. Near complete recovery of rCMRglc occurred by 7 days in young and old rats. We conclude that the basalocortical cholinergic projection plays a smaller role in neocortical function of aged rats, possibly because its tonic activity is reduced. Both young and aged rats undergo cortical metabolic normalization after unilateral NBM ablation; hence the NBM-lesioned aged rat is not a better model of the progressive decline in rCMRglc that occurs in Alzheimer's disease.

Electron-impact and chemical ionization detection of nicotine and cotinine by gas chromatography-mass spectrometry in rat plasma and brain.

Nicotine and its metabolite, cotinine, were measured in rat plasma and brain by gas chromatography-mass spectrometry. Both agents were extracted from plasma and brain, separated on a capillary column, and quantified by single-ion monitoring. The major fragment ions of nicotine and cotinine at m/z 84 and m/z 98, respectively, were monitored by electron-impact ionization detection and the protonated molecular ions at m/z 163 and m/z 177, respectively, were monitored by chemical ionization detection. Both compounds were quantified using deuterium-labeled nicotine and cotinine, respectively, as internal standards.

Adrenalectomy or metyrapone-pretreatment abolishes cerebral metabolic responses to the serotonin agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) in the hippocampus.

1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), a serotonin type 2 (5-HT2) agonist, elevates plasma corticosterone levels and reduces the cerebral metabolic rate for glucose (rCMRglc) in the hippocampus, a structure which possesses few 5-HT2 receptors but a large number of steroid receptors. To explore the hypothetical interaction between 5-HT and steroid mechanisms in the hippocampus, we measured rCMRglc in intact, adrenalectomized and metyrapone-pretreated rats after saline or DOI administration. Metyrapone pretreatment alone had no significant effect on rCMRglc, but adrenalectomy produced widespread rCMRglc increases in the cortex, hippocampus and monoaminergic brainstem nuclei. In intact rats, DOI 10 mg/kg reduced rCMRglc in limbic areas and increased it in the interanteromedial and paracentral thalamic nuclei. Metyrapone pretreatment and adrenalectomy abolished rCMRglc responses to DOI in hippocampal areas and enhanced those in thalamic nuclei. These results indicate that brain responses to DOI are dependent upon the functional state of the hypothalamus-pituitary-adrenal cortex axis.

Chronic treatment with meta-chlorophenylpiperazine (m-CPP) alters behavioral and cerebral metabolic responses to the serotonin agonists m-CPP and quipazine but not 8-hydroxy-2(di-N-propylamino)tetralin.

The effects of the serotonin (5-HT) agonists meta-chlorophenylpiperazine (m-CPP), quipazine and 8-hydroxy-2(di-n-propylamino)tetralin (DPAT) on behavior and on regional cerebral metabolic rates for glucose (rCMRglc) were measured in control rats or in rats pretreated for 2 weeks with continuous infusion of saline or m-CPP (2.5 mg/kg/day, subcutaneously). rCMRglc was measured in 71 brain regions, using the quantitative autoradiographic [14C]2-deoxy-D-glucose technique, at 15 min after acute administration of m-CPP 2.5 mg/kg, 60 min after quipazine 20 mg/kg, or 10 min after DPAT 1 mg/kg. Behavioral effects were assessed for m-CPP with an activity monitor, for quipazine by counting head shakes and for DPAT by scoring the serotonin syndrome. Chronic m-CPP pretreatment produced tolerance to hypolocomotion induced by acute m-CPP and to head shakes caused by acute quipazine, but did not alter the serotonin syndrome produced by DPAT. m-CPP 2.5 mg/kg IP produced widespread rCMRglc reductions in control rats but failed to modify rCMRglc in any region after chronic m-CPP pretreatment. Quipazine increased rCMRglc in 4 regions in control rats, but reduced rCMRglc in 14 brain areas of chronically m-CPP-pretreated animals. DPAT altered rCMRglc to the same degree in control (25 regions affected) and in chronically m-CPP-pretreated rats (28 regions affected). Reduced behavioral and metabolic effects of acute m-CPP in chronically m-CPP-pretreated rats were not due to pharmacokinetic alterations. These results demonstrate that chronic administration of m-CPP produces behavioral and metabolic tolerance to acute administration of m-CPP, but not of DPAT.(ABSTRACT TRUNCATED AT 250 WORDS)

Age-related alterations in behavioral and cerebral metabolic responses to the serotonin agonist meta-chlorophenylpiperazine in rats.

To determine the functional relevance of the age-related neurochemical changes that occur in brain serotonin systems during aging, we measured the effects of the serotonin receptor agonist meta-chlorophenylpiperazine (MCPP) on behavior and on regional cerebral metabolic rates for glucose (rCMRglc) in awake rats. rCMRglc was determined in 74 regions of Fischer-344 rats aged 3, 12 and 24 months, at 15 and 90 min after MCPP 2.5 mg/kg IP, using the quantitative, autoradiographic [14C]2-deoxy-D-glucose technique. The time-course of motor performance following MCPP was assessed with a rotating rod. MCPP impaired motor performance in all ages maximally at 15-30 min. Three-month-old rats recovered completely within 60 min, whereas 12-month-old animals exhibited partial recovery and 24-month-old rats did not recover by 120 min. At 15 min after MCPP, rCMRglc was reduced in 51 of the 74 studied regions (overall decrease, 20%) of 3-month-old rats, in 21 regions (13% decrease) of 12-month-old rats and in 14 regions (2% decrease) of 24-month-old animals. Similar MCPP brain concentrations were achieved at 15 min in rats of all ages. The results suggest that the functional integrity of serotonergic transmission is reduced in aged rats and that the dysregulation is presynaptic.

Parachloroamphetamine selectively alters regional cerebral metabolic responses to the serotonergic agonist metachlorophenylpiperazine in rats.

To determine if reported reductions of regional cerebral metabolic rates for glucose (rCMRglc) induced by the 5-HT agent metachlorophenylpiperazine (MCPP) (2.5 mg/kg) are due to a presynaptic action, 3-month old Fischer-344 rats were given parachloroamphetamine (PCA), a serotonin neurotoxin, and rCMRglc was measured 1 or 3 weeks later with the quantitative autoradiographic [14C]2-deoxyglucose procedure in 74 brain regions after administering saline, MCPP or other drugs. PCA alone increased rCMRglc significantly only in the raphe nuclei and in visual structures (visual cortex, lateral geniculate, superior colliculus). MCPP alone reduced rCMRglc in 75% of the regions studied. In PCA-lesioned rats, metabolic responses to MCPP 2.5 mg/kg were virtually abolished and rCMRglc was increased in interanteromedial and centrolateral thalamic nuclei. rCMRglc responses to quipazine, a postsynaptic serotonin agonist, and to arecoline and bromocriptine, cholinergic and dopaminergic agonists, were unchanged by PCA-pretreatment. Selective abolition by PCA of the metabolic response to MCPP confirms that MCPP, at the dose studied, reduces rCMRglc in the forebrain via a presynaptic mechanism and that postsynaptic serotonergic function is not altered by PCA.