Effect of donepezil in Alzheimer disease can be measured by a computerized human analog of the Morris water maze.

BACKGROUND: Drug development for Alzheimer disease (AD) is challenged by the success in animal models tested in the Morris water maze (MWM) and the subsequent failures to meet primary outcome measures in phase II or III clinical trials in patients. The human variant of MWM (hMWM) enables us to examine allocentric and egocentric navigation as in the MWM. OBJECTIVE: It was the aim of this study to examine the utility of a computerized hMWM to assess the effects of donepezil in mild AD. METHODS: Donepezil 5 mg/day was started after initial hMWM testing in the treated group (n = 12), and after 28 days, the dose was increased to 10 mg/day. The performance after 3 months was compared to that of a non-treated group (n = 12). RESULTS: Donepezil stabilized or improved the spatial navigation performance after 3 months, especially in the allocentric delayed recall subtask (p = 0.014). CONCLUSIONS: The computerized hMWM has the potential to measure the effects of donepezil in mild AD. It is a sensitive cognitive outcome measure in AD clinical trials.

Neuropeptide dietary supplement N-PEP-12 enhances cognitive function and activates brain bioelectrical activity in healthy elderly subjects.

N-PEP-12 is a dietary supplement consisting of neuropeptides and amino acids. In animal experiments, the compound has been shown to enhance cognitive function and reduce neurodegenerative events associated with aging. In this study, we investigated the effects of a single oral dose of N-PEP-12 (180 mg) on brain bioelectrical activity and cognitive performance in healthy elderly subjects. N-PEP-12 induced a significant (p < 0.05) increase in relative alpha-activity power 6 h after administration. This enhancement was accompanied by a generalized decrease in slow Delta-activity. Significant improvement in memory performance subtests was also seen 6 h after N-PEP-12 administration in some but not in all tests. Taken together, these data suggest that N-PEP-12 might be a reliable dietary supplement to be investigated for improving and, perhaps, maintaining brain function among healthy older adults.

Effects of Cerebrolysin on the outgrowth and protection of processes of cultured brain neurons.

Cerebrolysin (Cere, EBEWE Arzneimittel, Austria), a peptidergic drug produced by a standardised enzymatic breakdown of porcine brain proteins, consists of a mixture of 75% free amino acids and 25% low molecular weight peptides (<10 k DA). Cerebrolysin was shown to protect against MAP2 loss in primary embryonic chick neuronal cultures after brief histotoxic hypoxia and in a rat model of acute brain ischemia. Since MAP2 is involved in processes like neuronal growth, plasticity and dendritic branching, we address the question whether Cere is protecting processes against degeneration in a chronic low serum (2% FCS) cell stress model and whether the spontaneous outgrowth of axon-like processes is influenced. This was accomplished by quantification of the neurite lengths of embryonic chicken telencephalon neurons after 4 and 8 days. Additionally, time-lapse video microscopy was performed to study a possible influence of Cere on the growth cone behaviour of axon-like processes. To distinguish between effects caused by the peptide fraction and the effects related to free amino acids, we used an artificial amino acid solution (AA-mix). Results demonstrate a process outgrowth promoting effect of the AA-mix and Cere after 4 DIV. After 8 days neuronal network degeneration occurred in the AA-mix treated cultures, whereas Cere treated cultures still presented a well differentiated neuronal network. Dying neurons could release factors possibly impeding neurite outgrowth and Cere was shown to increase the viability of chicken cortical neurons. Neither the addition of BDNF nor serum supplementation (5% and 10% FCS) could protect the neuronal network against degeneration after 8 DIV, although these treatments were shown to ameliorate the viability of chicken telencephalon neurons. This result together with the finding obtained using the artificial amino acid solution points to the peptide fraction of Cere to be responsible for the protection of processes against degeneration. Time-lapse studies of Cere treated cultures revealed a significant decrease of the velocities characterising random growth cone movements, which is thought to be responsible for an increase in the length of axon-like processes after 4 DIV.

Antiapoptotic effects of the peptidergic drug cerebrolysin on primary cultures of embryonic chick cortical neurons.

Cerebrolysin (EBEWE Arzneimittel, Austria, Europe) is a widely used drug relieving the symptoms of a variety of neurological disorders, particularly of neurodegenerative dementia of the Alzheimer's type. It consists of approximately 25% of low molecular weight peptides (<10k DA) and a mixture of approximately 75% free amino acids, this being based on the total nitrogen content. In this study we used a low serum (2% serum supplement) cell stress in-vitro model to assess drug effectiveness on neuronal viability and programmed cell death (PCD). In this in-vitro model the type of cell death was previously shown to be primarly apoptotic, which was verified by DNA-laddering and TUNEL-staining. For evaluation of neuronal viability a MTT-reduction assay was performed after 4 DIV and 8 DIV and the percentage of apoptotic neurons was determined by bis-benzimide staining of nuclear chromatin. To differentiate between possible effects of the free amino acids and the peptide fraction of Cerebrolysin an artificial amino acid mixture (AA-mix) was used as a control. Cerebrolysin, the AA-mix and 10% foetal calf serum (FCS) caused a similar increase in viability after 4 DIV, whereas the effects of the growth factors BDNF and FGF-2 were less pronounced. After 8 DIV Cerebrolysin, but not the AA-mix, was able to ameliorate neuronal viability, which could reflect a neuro-protective effect or an increased activity of the mitochondrial dehydrogenase measured in a MTT-reduction assay. The percentage of cells showing apoptotic chromatin changes was significantly reduced (p < 0.01) in cultures treated with Cerebrolysin, whereas the AA-mix failed to decrease the percentage of cells showing apoptotic chromatin changes. These findings ascertain an anti-apoptotic effect of the peptide fraction of Cerebrolysin and reveal a transient viability promoting effect of the amino acid fraction, which is most likely due to improved nutritional supply.

A brain derived peptide preparation reduces the translation dependent loss of a cytoskeletal protein in primary cultured chicken neurons.

Neuronal cytoskeletal proteins like the microtubule associated protein 2 (MAP2) are objected to pathological proteolysis in case of Alzheimer's disease and brain ischemia. The neurotrophic peptidergic drug Cerebrolysin (EBEWE Arzneimittel, Austria, Europe) is produced by a standardized enzymatic break-down of lipid free porcine brain proteins. Cerebolysin protected MAP2 in primary neuronal cultures after a brief histotoxic hypoxia and in a rat model of acute brain ischemia. Furthermore the drug was shown to inhibit the proteases mu- and m-calpain dose dependently in several cell free protease activity assays. The question if the higher MAP2 levels are due to an alleviation of proteolysis, to a higher synthesis rate or both is addressed in the current investigation: Monitoring the MAP2 content of primary neuronal cell cultures over a period of eight days revealed MAP2 to reach a peak level on day six in vitro followed by a degradation phase. In other experiments the protein synthesis of Cerebrolysin treated and untreated cells was blocked with cycloheximide at that moment when all cells exhibited the same MAP2 content. After the following MAP2 degradation phase--i.e. after eight days in vitro--the MAP2 contents were determined by western blotting. Cerebrolysin treated cells contained more MAP2 than untreated controls proving that the drug protects MAP2 independently from de novo synthesis, although further work is in progress to investigate if the drug supplementary boosts this effect by increasing MAP2 synthesis.

Ameliorative influence of a nootropic drug on motor activity of rats after bilateral carotid artery occlusion.

The effects of the peptidergic nootropic drug Cerebrolysin on spatial memory and motor activity were examined in intact and ischemic rats. Ischemic-hypoxic damage was induced by injection of Na-cyanide followed by bilateral occlusion of common carotid arteries. Immediately afterwards Cerebrolysin or saline was administered, either by continuous intraventricular (i.v.) infusion or by daily intraperitoneal (i.p.) injection. Rats were tested for spatial memory and motor activity in the Morris water maze on days 3 and 4 post-surgery. The best dose of the substance for i.p. administration was known from previous studies. Therefore we had to investigate the dose-response-relationship and tolerability of the drug after i.v. administration in intact rats. Infusion (i.v.) of a high dose of Cerebrolysin (0.57 mg/day) decreased motor activity and spatial memory of intact rats (p < 0.01 and p < 0.05, respectively) but low dose of Cerebrolysin was well tolerated in the intact animals. Ischemia led to deterioration of motor activity in control rats (p < 0.01). Cerebrolysin significantly counteracted deleterious motor changes due to ischemia up to the level of intact controls after both i.v. infusion (0.0057 mg/day) and daily i.p. drug administration (100 mg/kg bw and day) indicating an accelerating recovery after ischemia.