![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
NEUROPHARMACOLOGY
Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, Chicago, Illinois
Received February 25, 2004; accepted April 29, 2004.
 |
Abstract |
|---|
 Top Abstract Materials and MethodsResultsDiscussionReferences |
|---|
-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid and kainate currents were not affected by galantamine. The maximum potentiation of NMDA currents to 
130% of the control was obtained at 1 µM galantamine. The potentiation was due to a shift of the NMDA dose-response curve in the direction of lower NMDA concentrations. Glycine at 1 to 3000 nM enhanced NMDA currents, and potentiation by 1 µM galantamine and 1 to 300 nM glycine was additive. The glycine site antagonist 7-chlorokynurenic acid did not prevent the galantamine action. These results suggested that galantamine did not interact with the glycine binding site. Experiments with various concentrations of Mg2+ indicated that galantamine did not affect the Mg2+ blocking site of the NMDA receptor. PKC was involved in galantamine potentiation of NMDA currents, but protein kinase A, Gi/Go proteins, and Gs proteins were not involved. Potentiation of the activity of NMDA receptors is deemed partially responsible for the improvement of cognition, learning, and memory in Alzheimer's patients.
; Woodruff-Pak and Hinchliffe, 1997), one of the strategies for improving the cognitive activity of the patients would be to stimulate nAChRs. This can be accomplished by inhibiting acetylcholinesterase. In fact, four of five Alzheimer's drugs that have been approved by the Food and Drug Administration for clinical use in the United States, tacrine, donepezil, rivastigmine, and galantamine, are anticholinesterases.
Clinical tests with Alzheimer's patients and animal experiments have indicated the effectiveness of galantamine to improve cognition (Kroll et al., 1999; Barnes et al., 2000; Raskind et al., 2000; Tariot et al., 2000). Galantamine has been shown to act as an agonist generating single-channel currents in -bungarotoxin-insensitive, 4
2-type nAChRs of cultured rat hippocampal neurons (Pereira et al., 1993) and in the chicken 42 receptors expressed in fibroblast (M10) cells (Pereira et al., 1994). However, galantamine failed to generate a detectable whole-cell current in M10 cells expressing the 42 receptors (Pereira et al., 1994). It potentiated ACh-induced whole-cell current in PC12 cells (3-type AChRs), in hippocampal 7-type AChRs (Schrattenholz et al., 1996; Maelicke and Albuquerque, 2000), in 42 AChRs expressed in human embryonic kidney cells (Samochocki et al., 2003), and in 7-type AChRs in 7/5-HT3 chimera (Maelicke et al., 2001). Galantamine was also found to potentiate nicotine-evoked increases in intracellular Ca2+ and [3H]noradrenaline release in SH-SY5Y cells (Dajas-Bailador et al., 2003). The potentiating action of galantamine on nicotinic receptors facilitated synaptic transmission in the brain (Santos et al., 2002).
In addition to nAChRs, glutamate receptors are also impaired in the brain of Alzheimer's patients (Palmer, 2002). Changes in the glutamatergic system are known to cause cognitive impairments. For instance, the loss of perikarya of piramidal cells and of the midtemporal gyrus was found to correlate with the severity of dementia (Palmer and Gershon, 1990). In Alzheimer's patients, glutamate-containing neurons are decreased (Greenamyre et al., 1987; Palmer and Gershon, 1990; Francis et al., 1993; Palmer, 1996). Memantine is a low-affinity noncompetitive antagonist of the N-methyl-D-aspartate (NMDA) receptor and has recently been approved by the Food and Drug Administration for treating moderate-to-severe Alzheimer's disease patients (Tariot et al., 2004).
Drugs that modulate the NMDA receptor-mediated neural transmission by acting at the glycine site are potential therapeutic agents to treat memory deficits associated with aging and Alzheimer's disease. Both the partial glycine site agonist d-cycloserine and the glycine prodrug milacemide facilitate memory in animal paradigms (Hanndelmann et al., 1989; Baxter et al., 1994) and have been tested as cognitive enhancers in patients with Alzheimer's disease (Schwartz et al., 1991, 1996; Dysken et al., 1992).
Because NMDA receptors seem to play a crucial role in learning and memory, our working hypothesis is that one of the mechanisms of action of galantamine is to modulate the NMDA receptor functions. Thus, the cognitive function of patients with Alzheimer's disease or with other forms of dementia who have reduced NMDA receptors may be improved by increasing the activity of NMDA receptors. We have indeed found in the present study that galantamine augments NMDA-evoked currents in rat cortical neurons. This action seems to be exerted via the PKC system.
|
Materials and Methods |
|---|
|
TopAbstractMaterials and MethodsResultsDiscussionReferences |
|---|
). In brief, rat embryos were removed from a 17-day pregnant Sprague-Dawley rat under halothane anesthesia. Small wedges of frontal cortex were excised and subsequently incubated in phosphate buffer solution for 20 min at 37°C. This solution contained 154 mM NaCl, 1.05 mM KH2PO4, 3.0 mM Na2HPO4·7 H2O, 0.25% (w/v) trypsin (type XI; Sigma-Aldrich, St. Louis, MO), pH 7.4, and with an osmolarity of 287 mOsM. The digested tissue was then mechanically triturated by repeated passages through a Pasteur pipette, and the dissociated cells were suspended in neurobasal medium with B-27 supplement (Invitrogen, Carlsbad, CA) and 2 mM glutamine. The cells were added to 35-mm culture wells at a concentration of 100,000 cells/ml. Each well contained five 12-mm poly-L-lysine-coated coverslips overlaid with confluent glia that had been plated 2 to 4 weeks earlier. The cortical neuron/glia cultures were maintained in a humidified atmosphere of 90% air and 10% CO2 at 34°C. Cells cultured for 3 to 7 weeks were used for experiments. Solutions for Current Recording. The external solution for recording of the whole-cell currents contained 150 mM NaCl, 5 mM KCl, 2.5 mM CaCl2, 1 mM MgCl2, 5.5 mM HEPES acid, 4.5 mM HEPES sodium, 10 mM D-glucose, pH 7.3, and the osmolarity was adjusted to 300 mOsM with D-glucose. Tetrodotoxin (100 nM) was added to eliminate the voltage-gated sodium channel currents. Atropine sulfate (20 nM) was added to block the muscurine AChR currents. The internal pipette solution contained 140 mM potassium-gluconate, 2 mM MgCl2, 1 mM CaCl2, 10 mM HEPES acid, 10 mM EGTA, 2 mM ATP-Mg2+, and 0.2 mM GTP-Na+. The pH was adjusted to 7.3 with KOH and the osmolality was adjusted to 300 mOsM by adding D-glucose.
Whole-Cell Current Recording. Ionic currents were recorded using the whole-cell patch-clamp technique at room temperature (21–22°C). Pipette electrodes were made from 1.5-mm (outer diameter) borosilicate glass capillary tubes with a resistance of 2 to 3 M
when filled with the standard internal solution. The membrane potential was clamped at -70 mV unless otherwise stated. We allowed 5 to 10 min after membrane rupture for the cell interior to adequately equilibrate with the pipette solution. Currents through the electrode were recorded with an Axopatch-1C amplifier (Axon Instruments Inc., Union City, CA), filtered at 2 kHz, and sampled at 10 kHz in a PC-based data acquisition system that also provided preliminary data analysis. Results are expressed as mean ± S.E.M., and n represents the number of the cells examined.
Drug Applications. Two methods for drug application were used: one was perfusion through the bath and the other was via a U-tube controlled by a computer-operated magnetic valve system (Marszalec and Narahashi, 1993). In the fast U-tube application system, when one of the valves was open, the drug solution was allowed to bypass the chamber. When it was closed, the drug solution was ejected through the hole of the U-tube, which was located close to the cell. At the same time, another valve controlling the suction tube was opened, allowing a laminated flow of the test solution across the recording cell such that the external solution surrounding the cell could be completely changed with the drug solution within 30 to 40 ms. In some cases, test drugs were also added to the external solution and continuously perfused to the recording chamber via a glass syringe and Teflon tube.
Chemicals. NMDA (Sigma-Aldrich), AMPA(Sigma/RBI, Natick, MA), kainate (Sigma/RBI), glycine (Sigma/RBI), and 7-chlorokynurenic acid (7-ClKN) (Sigma/RBI) were first dissolved in distilled water to make stock solutions. The Gi/Go protein inhibitor pertussis toxin, the Gs protein stimulator cholera toxin, the voltagegated sodium channel blocker tetrodotoxin, the muscarine AChR blocker atropine sulfate, and the PKC inhibitor chelerythrine chloride were purchased from Sigma-Aldrich. The PKA inhibitor H-89 was obtained from Calbiochem-Novabiochem (La Jolla, CA). Galantamine was a gift from Janssen Pharmaceutica (Titusville, NJ) and was first dissolved in distilled water to make stock solutions. The stock solutions were stored at 4°C and diluted to prepare test solutions with the standard external solution shortly before the experiments.
Data Analyses. Current records were initially analyzed by the Clamp-Fit module of the PClamp6 program to assess the whole-cell peak current amplitude and decay kinetics. Data were expressed as the mean ± S.E.M. unless otherwise stated. The concentration-response data were subsequently compiled for graphical analysis in SigmaPlot 8.0. Analysis of variance and Student's t tests were performed to assess the significance of differences, and the p values less than 0.05 were considered statistically significant.
|
Results |
|---|
|
TopAbstractMaterials and MethodsResultsDiscussionReferences |
|---|
|
Dose-response relationships for galantamine potentiation of NMDA currents in multipolar neurons are shown in Fig. 1, C and D. The minimal effective concentration of galantamine to potentiate NMDA currents was 10 nM at which only about 5% current potentiation over and above the control was observed. Galantamine at 1 µM potentiated the current by about 30%, yet a further increase in the concentration to 10 µM caused a decrease in efficacy. This resulted in a bell-shaped dose-response relationship with a maximal efficacy at 1 µM galantamine (Fig. 1D). The bell-shaped dose-response curve was also observed in nefiracetam potentiation of the 42 AChR currents (Zhao et al., 2001) and the NMDA receptor currents (Moriguchi et al., 2003a), and animal behavioral responses to nefiracetam (Nabeshima et al., 1994).
At least two potential mechanisms of galantamine potentiation of NMDA currents are conceivable. One is an increase in the affinity of the receptor for NMDA, which would reflect in a shift of the NMDA dose-response curve in the direction of lower concentrations of NMDA. The other possibility is an increase in current amplitude even at high concentrations of NMDA that produce a saturating response without changing the NMDA affinity for the binding site. Dose-response relationships for NMDA to induce currents before and during bath and U-tube applications of 1 µM galantamine are illustrated in Fig. 2A. The peak current amplitudes normalized to the maximum control current induced by 1000 µM NMDA were fitted by a sigmoid curve with an EC50 value of 36.7 ± 0.2 µM and a Hill coefficient of 0.60 ± 0.09 (Fig. 2A, 
; n = 4). After a 10-min bath and U-tube applications of 1 µM galantamine, the maximal current amplitude induced by 1000 µM NMDA was 104.8 ± 6.8% of the control maximum, an EC50 was 26.2 ± 0.4 µM, and a Hill coefficient was 0.64 ± 0.07 (Fig. 2A, 
; n = 4). Thus, the dose-response curve was shifted significantly (p < 0.05) in the direction of lower NMDA concentrations by galantamine application without changing the amplitude of maximum current, indicating that galantamine increased the receptor affinity for NMDA. This resulted in a decrease in the percentage of galantamine potentiation of NMDA currents with an increase in NMDA concentration (Fig. 2B). The situation is in sharp contrast with nefiracetam potentiation of the 42 AChR currents (Zhao et al., 2001) and NMDA receptor currents (Moriguchi et al., 2003a), in which currents were increased beyond the saturating response produced by high concentrations of ligands.
|
Does Galantamine Interact with the Glycine Binding Site of NMDA Receptors? Our previous study showed that nefiracetam interacted with the glycine binding site of the NMDA receptors (Moriguchi et al., 2003a). The interactions between glycine and galantamine were studied using various concentrations of glycine on the NMDA currents induced by 30 or 300 µM NMDA (Fig. 3). Figure 3A shows that glycine, when applied via the bath and U-tube, potentiated NMDA currents in a concentration-dependent manner until the current amplitude reached a maximum at 3000 nM glycine. Similar glycine potentiation was also observed in the presence of 1 µM galantamine in the bath and U-tube (Fig. 3A). Galantamine potentiation was 29% in nominal glycine-free solution and those in the presence of 10, 30, 100, and 300 nM glycine were 30, 37, 31, and 26%, respectively. Thus, galantamine and glycine act additively in potentiating NMDA currents except at very high concentrations of glycine (1000 and 3000 nM). When the NMDA currents were induced by 300 µM NMDA, glycine still potentiated the NMDA current (Fig. 3B), but galantamine failed to further enhance the current. Thus, it seems that galantamine does not interact with the glycine binding site of the NMDA receptor.
|
To further corroborate the aforementioned notion, galantamine potentiation of NMDA currents was tested in the presence of 1 µM 7-ClKN, a specific inhibitor of the glycine binding site of the NMDA receptor. 7-ClKN drastically suppressed NMDA currents, yet 1 µM galantamine was able to potentiate the currents to 149 ± 6.0% of the control (p < 0.0002; n = 4) in the presence of 7-ClKN (Fig. 4). In keeping with this result, the dose-response curve of the 7-ClKN inhibition of NMDA currents was not affected by the presence of 1 µM galantamine (Fig. 5). The IC50 values estimated from the dose-response relationships are not significantly different (p > 0.05), being 298 ± 26.5 and 316.6 ± 24.6 nM in the absence and presence of galantamine, respectively. Thus, it was concluded that galantamine did not interact with the glycine binding site of the NMDA receptor.
|
|
Does Galantamine Interact with the Mg2+ Binding Site of NMDA Receptors? Mg2+ ions are known to block the NMDA receptor in a voltage-dependent manner, especially at large negative membrane potentials. Partial block occurs at the normal resting potential. Thus, our hypothesis is that galantamine potentiates NMDA currents by eliminating Mg2+ block. This hypothesis was tested by the following experiments.
When the membrane potential was held at -70 mV, Mg2+ inhibited NMDA currents in a concentration-dependent manner, almost completely blocking the currents at 1000 µM (Fig. 6A). Galantamine at 1 µM potentiated NMDA currents, and Mg2+ concentrations ranging from 1 to 1000 µM also suppressed the currents (Fig. 6B). The dose-response curves for Mg2+ block without and with 1 µM galantamine are plotted in Fig. 6C. The IC50 for Mg2+ to block the NMDA current was 121 ± 9.6 µM in the absence of galantamine. Galantamine increased the NMDA current by 15, 16, 17, 15.4, and 28% in the presence of 0, 1, 10, 100, and 1000 µM Mg2+, respectively. An addition of 16% to the NMDA currents predicted from the dose-response relationship in the absence of galantamine could fit the data obtained in the presence of galantamine with the exception of a very high concentrations of Mg2+ at 1000 µM.
|
The weaker suppression of NMDA currents by 1 mM Mg2+ in the presence of galantamine was further investigated in the experiments shown in Fig. 7. In addition to blocking the channel, external millimolar Mg2+ ions have also been shown to potentiate NMDA currents under certain conditions (Paoletti et al., 1995; Wang and MacDonald, 1995). To assess the effect of galantamine on the Mg2+ potentiation of the NMDA current, the current was recorded at +40 mV in the presence of 1 mM Mg2+ and 30 nM glycine. Mg2+ at 1 mM alone did not change the NMDA current (date not shown), but increased 13.2 ± 2.3% (n = 5) over and beyond the control level in the presence of 1 µM galantamine. With this correction, the galantamine-induced increase of the NMDA current in the presence of 1 mM Mg2+ at -70 mV became 15%, which is in line with the increases in the absence of Mg2+ ions or in lower Mg2+ concentrations. Thus, galantamine did not alter Mg2+ block of the NMDA receptor even in the presence of 1 mM Mg2+.
|
Role of PKA, PKC, and G Proteins in Galantamine Potentiation. Nefiracetam potentiates the activity of the 42 nAChR via Gs proteins, but not Gi/Go proteins, PKA, or PKC (Zhao et al., 2001). To assess the roles of these G proteins and kinases in galantamine potentiation of NMDA currents, the following experiments were conducted using specific chemicals agents.
The membrane-permeable PKC-specific inhibitor chelerythrine, when applied to the bathing solution at a concentration of 3 µM, slightly suppressed NMDA currents (Fig. 8, A and B). This is in keeping with the observation that the activation of PKC potentiates NMDA responses (Chen and Huang, 1991; Ben-Ari et al., 1992; Raymond et al., 1994; Leonard and Hell, 1997; Logan et al., 1999). Bath and U-tube applications of 1 µM galantamine in the presence of chelerythrine did not potentiate NMDA currents (Fig. 8, A and B). In the presence of 3 µM PKC inhibitor chelerythrine, 1 µM galantamine did not significantly increase the NMDA current (104.0 ± 4.6% of the control; p > 0.5). Thus, PKC is involved in the galantamine potentiation of NMDA currents.
|
The membrane-permeable PKA-specific inhibitor H-89, when applied to the bath at a concentration of 1 µM, slightly suppressed NMDA currents (Fig. 8, C and D). Unlike PKC inhibition, 1 µM galantamine was still able to potentiate NMDA currents 117.2 ± 6.20% of the control (p < 0.04) (Fig. 8, C and D). Thus, PKA is not involved in the galantamine potentiation of NMDA currents.
Potentiation of NMDA currents by 1 µM galantamine was not prevented by 24- to 26-h preincubation of neurons with either 200 ng/ml pertussis toxin, a Gi/Go protein inhibitor, or 500 ng/ml cholera toxin, a Gs protein stimulator (Fig. 9). The currents were increased by galantamine to 123.8 ± 3.7% (p < 0.0003) of the control after pertussis toxin treatment and to 117.8 ± 3.4% (p < 0.002) after cholera toxin treatment. Thus, Gi/Go and Gs proteins are not involved in the galantamine potentiation of NMDA currents.
|
AMPA and Kainate Currents Are Not Affected by Galantamine. Besides the NMDA currents, rat cortical neurons in primary culture generated currents in response to U-tube application of AMPA or kainate. Our previous study showed that nefiracetam did not significantly potentiate APMA- and kainate-induced currents (Moriguchi et al., 2003a). Currents induced by 30 µM AMPA (n = 4) or 30 µM kainate (n = 4) in multipolar neurons were not changed by bath and U-tube applications of 1 µM galantamine (Fig. 10). The lack of effect of galantamine on the AMPA-induced current, which is mediated by the AMPA receptors, and the lack of effect of galantamine on the kainate-activated current, which could be mediated by both AMPA and kainate receptors, led us to conclude that galantamine has no potentiating action on both receptors. Thus, it was concluded that among the three subtypes of glutamate receptors, the AMPA and kainate receptors were not responsive to the modulating action of galantamine, and only the NMDA receptors responded to galantamine.
|
|
Discussion |
|---|
|
TopAbstractMaterials and MethodsResultsDiscussionReferences |
|---|
Alzheimer's disease is a progressive neurodegenerative disorder of cognitive function. It is known that the Alzheimer's disease is associated with down-regulation of both nAChRs and NMDARs in the brain (Fonnum et al., 1995; Giacobini, 2000). Thus, potentiation of the activity of these systems is expected to improve learning/memory/cognition of Alzheimer's patients. Galantamine potentiation of NMDA currents described here is deemed to contribute significantly to the therapeutic effects.
Galantamine weakly inhibits acetylcholinesterase with an IC50 value of 600 to 800 nM. However, galantamine directly stimulates nAChRs also. Earlier studies showed that galantamine at 1 to 10 µM opened single AChR channels in the rat hippocampus and in the chicken 42 receptors expressed in mouse fibroblast (M10) cells but failed to evoke measurable whole-cell currents (Pereira et al., 1993, 1994). However, acetylcholine-induced whole-cell currents were potentiated by galantamine (0.1–1 µM) in PC12 cells (3-type AChRs), in hippocampal 7-type AChRs (Schrattenholz et al., 1996; Maelicke and Albuquerque, 2000), in 42 AChRs expressed in human embryonic kidney cells (Samochocki et al., 2003), and in 7/5HT3 chimera (Maelicke et al., 2001). Because galantamine potentiation was blocked by the monoclonal antibody FK1 that recognizes the subunit of nAChRs, galantamine is thought to bind to a site distinct from the ACh binding site (Schrattenholz et al., 1996) acting as an allosterically potentiating ligand. A bell-shaped dose-response relationship was obtained with the potentiation peaking at 1 µM (Schrattenholz et al., 1996), but the underlying mechanism has yet to be elucidated.
We have already reported that nefiracetam potentiates NMDA currents in the nominal absence of glycine or in the presence of low concentration of glycine (Moriguchi et al., 2003b). In fact, d-cycloserine, a glycine site partial agonist, is known to improve implicit memory performance of words in Alzheimer's patients (Schwartz et al., 1996) and reversal learning of rats in the water maze (Riekkinen et al., 1998). Thus, nefiracetam and d-cycloserine, as the glycine site partial agonists, share the common mechanism of action with respect to stimulation of NMDA receptors. In contrast, galantamine has a different mechanism of action from nefiracetam in potentiating NMDA currents (Moriguchi et al., 2003a).
Mg2+ ions at lower concentrations are known to block the NMDA channel in a voltage-dependent manner with the block augmented at negative potentials and attenuated at positive potentials (Mayer et al., 1984; Ascher and Nowak, 1988). Galantamine does not seem to interact with the Mg2+ blocking site of the NMDA receptor. Mg2+ ions at higher concentrations are known to enhance the NMDA currents, which is best seen at positive potentials. Galantamine enhanced the NMDA current measured at +40 mV in the presence of 1 mM Mg2+ ions. The latter observation might explain some of the differences between galantamine and nefiracetam on the NMDA current. A reduction in Mg2+ block was observed in the presence of nefiracetam (Moriguchi et al., 2003b) but not in the presence of galantamine.
It is well established that protein phosphorylation plays an important role in various neuroreceptors (Huganir and Greengard, 1990; Hoffman et al., 1994). Our experiments with the cortical neuron NMDA receptors indicated that PKC inhibition prevented galantamine potentiation but PKA inhibition did not. Pretreatment with the Gi/Go inhibitor pertussis toxin and the Gs stimulator cholera toxin did not prevent galantamine from potentiating NMDA currents. Thus, PKC seems to be involved in galantamine potentiation of NMDA currents.
Intracellular protein kinases are known to potentiate NMDA-mediated responses. Specifically, the activation of PKC is known to potentiate NMDA responses (Chen and Huang, 1991; Ben-Ari et al., 1992; Kelso et al., 1992; Urushihara et al., 1992; Logan et al., 1999) and to phosphorylate the receptor (Raymond et al., 1994; Leonard and Hell, 1997). PKC potentiation has been attributed to an increase in the channel open probability (Chen and Huang, 1992; Xiong et al., 1998) or a reduction in Mg2+ block (Chen and Huang, 1992). PKC activation has also been shown to reduce NMDA responses recorded from hippocampal CA1 pyramilal neurons (Markram and Segal, 1992) and in cerebellar granule cells (Snell et al., 1994). These differences in PKC modulation of NMDA responses might arise from different PKC isozymes (Nishizuka, 1988) and different subunit compositions of the NMDA receptors that exist in the brain (Snell et al., 1994).
The NMDA current was potently and efficaciously enhanced by nefiracetam via the PKC pathway (Moriguchi et al., 2003b). In the presence of nefiracetam, the Mg2+ block observed at -70 mV was almost completely eliminated. In the present study, the PKC inhibitor chelerythrine prevented the potentiating action of galantamine. Despite the involvement in the PKC pathway, the Mg2+ block was not affected by galantamine. The difference in Mg2+ block between nefiracetam and galantamine might be partly due to the degree of potentiation. Nefiracetam increased the NMDA current to nearly 300% of the control, whereas galantamine increased only by 30%. In addition, PKC activation has been shown to yield variable results on Mg2+ block, reducing Mg2+ block in some studies (Chen and Huang, 1991, 1992) but not in others (Wagner and Leonard, 1996).
Galantamine seems to work via multiple pathways to improve the memory/learning/cognition of Alzheimer's patients. The present study has demonstrated galantamine potentiation of the NMDA receptor activity. In addition to weakly inhibiting cholinesterase (Thomson et al., 1991), galantamine has been shown to alloterically potentiate the activity of nACh receptors (Pereira et al., 1993, 1994; Schrattenholz et al., 1996; Maelicke and Albuquerque, 2000; Samochocki et al., 2003). In the brain of Alzheimer's patients, not only nAChR activity is down-regulated (Vidal and Changeux, 1996; Woodruff-Pak and Hinchliffe, 1997) but also the glutamatergic system is impaired (Greenamyre et al., 1987; Palmer and Gershon, 1990; Francis et al., 1993; Palmer, 1996). Thus, galantamine stimulation of both nACh and glutamate systems is deemed to work in concert contributing to the improvement of the patient's conditions.
In conclusion, the Alzheimer's drug galantamine enhances the NMDA responses of rat cortical neurons in long-term primary culture. The potentiation of the NMDA receptor activity by galantamine is mediated by PKC but not by PKA or G proteins. The potentiation of the activity of NMDA receptors is an important aspect of galantamine action in improving learning/memory/cognition when the NMDA receptor activity is low in Alzheimer's patients.
|
Acknowledgements |
|---|
|
Footnotes |
|---|
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
ABBREVIATIONS: nAChR; nicotinic acetylcholine receptor; NMDA, N-methyl-D-aspartate; PKC, protein kinase C; AMPA, -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; 7-ClKN, 7-chlorokynurenic acid; PKA, protein kinase A; H-89, N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide hydrochloride.
Address correspondence to: Dr. Toshio Narahashi, Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, 303 East Chicago Ave., Chicago, IL 60611. E-mail: narahashi{at}northwestern.edu
|
References |
|---|
|
TopAbstractMaterials and MethodsResultsDiscussionReferences |
|---|
Ascher P and Nowak L (1988) The role of divalent cation in the N-methyl-D-aspartate responses of mouse central neurones in culture. J Physiol (Lond) 399: 247-266.
Barnes CA, Meltzer J, Houston F, Orr G, McGann K, and Wenk GL (2000) Chronic treatment of old rats with donepezil or galantamine: effects on memory, hippocampal plasticity and nicotinic receptors. Neuroscience 99: 17-23.[CrossRef][Medline]
Baxter MG, Lanthorn TH, Frink KM, Golski S, Wan R, and Olton DS (1994) d-Cycloserine, a novel cognitive enhancer, improves spatial memory in aged rats. Neurobiol Aging 15: 207-213.[CrossRef][Medline]
Ben-Ari Y, Aniksztein L, and Bregestovski P (1992) Protein kinase C modulation of NMDA currents: an important for LTP induction. Trends Neurosci 15: 333-339.[CrossRef][Medline]
Chen L and Huang LY (1991) Sustained potentiation of NMDA receptor-mediated glutamate responses through activation of protein kinase C by a mu opioid. Neuron 7: 319-326.[CrossRef][Medline]
Chen L and Huang LY (1992) Pritein kinase C reduces Mg2+ block of NMDA-receptor channels as a mechanism of modulation. Nature (Lond) 356: 521-523.[CrossRef][Medline]
Dajas-Bailador FA, Heimala K, and Wonnacott S (2003) The allosteric potentiation of nicotinic acetylcholine receptors by galantamine is transduced into cellular responses in neurons: Ca2+ signals and neurotoransmitter release. Mol Pharmacol 64: 1217-1226.
Dysken MW, Mendels J, and LeWitt P (1992) Milacemide; a placebo-controlled study in senile dementia of the Alzheimer type. J Am Geriatr Soc 40: 503-506.[Medline]
Fonnum F, Myhrer T, Paulsen RE, Wangen K, and Oksengard AR (1995) Role of glutamate and glutamate receptors in memory function and Alzheimer's disease. Ann NY Acad Sci 757: 475-486.[CrossRef][Medline]
Francis PT, Sims NR, Procter AW, and Bowen DM (1993) Cortical pyramidai neurone loss may cause glutamatergic hypoactivity and cognitive impairment in Alzheimer's disease: investigative and therapeutic perspectives. J Neurochem 60: 1589-1604.[CrossRef][Medline]
Giacobini E (2000) Cholinesterase inhibitor therapy stabilizes symptoms of Alzheimer's disease. Alzheimer Dis Assoc Disorders 14 (Suppl 1): S3-S10.[CrossRef]
Greenamyre JT, Penney JB, D'Amato CJ, and Young AB (1987) Dementia of the Alzheimer's type: changes in hippocampal L-[3H]glutamate binding. J Neurochem 48: 543-551.[CrossRef][Medline]
Hanndelmann GE, Nevins ME, Mueller LL, Arnolde SM, and Cordi AA (1989) Milacemide, a glycine prodrug, enhances performance of learning tasks in normal and amnestic rodents. Pharmacol Biochem Behav 34: 823-828.[CrossRef][Medline]
Hoffman PW, Ravindran A, and Hunganir RL (1994) Role of phosphorylation in desensitization of acetylcholine receptors expressed in Xenopus oocytes. J Neurosci 14: 4185-4195.[Abstract]
Huganir RL and Greengard P (1990) Regulation of neurotransmitter receptor desensitization by protein phosphorylation. Neuron 5: 555-567.[CrossRef][Medline]
Kelso SR, Nelson TE, and Leonard JP (1992) Protein kinase C-mediated enhancement of NMDA currents by metabotropic glutamate receptors in Xenopus oocytes. J Physiol (Lond) 449: 705-718.
Kroll WJ, Sramek JJ, and Cutler NR (1999) Cholinesterase inhibitors: a therapeutic strategy for Alzheimer disease. Ann Pharmacotherapy 33: 441-450.[Abstract]
Leonard AS and Hell JW (1997) Cyclic AMP-dependent protein kinase and protein kinase C phosphorylate N-methyl-D-aspartate receptors at different sites. J Biol Chem 272: 12107-12115.
Logan SM, Rivera FE, and Leonard JP (1999) Protein kinase C modulation of recombinant NMDA receptor currents: roles for the C-terminal C1 exon and calcium ions. J Neurosci 19: 974-986.
Maelicke A and Albuquerque EX (2000) Allosteric modulation of nicotinic acetylcholine receptors as a treatment strategy for Alzheimer's disease. Eur J Pharmacol 393: 165-170.[CrossRef][Medline]
Maelicke A, Samochocki M, Jostock R, Fehrenbacher A, Ludwig L, Albuquerque EX, and Zerlin M (2001) Allosteric sensitization of nicotinic receptors by galantamine, a new treatment strategy for Alzheimer's disease. Soc Biol Psychiatry 49: 279-288.
Markram H and Segal M (1992) Activation of protein kinase C suppresses responses to NMDA in rat CA1 hippocampal neurones. J Physiol (Lond) 457: 491-501.
Marszalec W and Narahashi T (1993) Use-dependent pentobarbital block of kainate and quiaqualate currents. Brain Res 608: 7-15.[CrossRef][Medline]
Mayer ML, Westbrook GL, and Guthrie PB (1984) Voltage dependent block by Mg2+ of NMDA responses in spinal cord neurones. Nature (Lond) 309: 251-263.
Moriguchi S, Marszalec W, Zhao X, Yeh JZ, and Narahashi T (2003a) Potentiation of N-methyl-D-aspartate-induced currents by the nootropic drug nefiracetam in rat cortical neurons. J Pharmacol Exp Ther 307: 160-167.
Moriguchi S, Marszalec W, Zhao X, Yeh JZ, and Narahashi T (2003b) Alzheimer's drug nefiracetam modulates the activity of NMDA receptors in rat cortical neurons. Soc Neurosci Abstr 795.12.
Nabeshima T, Nakayama S, Ichihara K, Yamada K, Shiotani T, and Hasegawa T (1994) Effects of nefiracetam on drug-induced impairment of latent learning in mice in a water finding task. Eur J Pharmacol 255: 57-65.[CrossRef][Medline]
Nishizuka Y (1988) The molecular heterogeneity of protein kinase C and its implications for cellular regulation. Nature (Lond) 334: 661-665.[CrossRef][Medline]
Palmer AM (1996) Neurochemical studies of Alzheimer's disease. Neurodegeneration 5: 381-391.[CrossRef][Medline]
Palmer AM (2002) Pharmacotherapy for Alzheimer's disease: progress and prospects. Trends Pharmacol Sci 23: 426-433.[CrossRef][Medline]
Palmer AM and Gershon S (1990) Is the neurochemical basis of Alzheimer's disease cholinergic or glutamatergic? FASEB J 4: 2745-2752.[Abstract]
Paoletti P, Neyton J, and Ascher P (1995) Glycine-independent and subunit-specific potentiation of NMDA responses by extracellular Mg2+. Neuron 15: 1109-1120.[CrossRef][Medline]
Pereira EFR, Alkondon M, Reinhardt S, Maelicke A, Peng X, Lindstrom P, Whiting P, and Albuquerque EX (1994) Physostigmine and galanthamine: probes for a novel binding site on the 4
2 subtypes of neuronal nicotinic acetylcholine receptors stably expressed in fibroblast cells. J Pharmacol Exp Ther 270: 768-778.
Pereira EFR, Reinhardt-Maelicke, Schrattenholz A, Maelicke A, and Albuquerque (1993) Identification and functional characterization of a new agonist site on nicotinic acetylcholine receptors in cultured hippocampal neurons. J Pharmacol Exp Ther 265: 1474-1491.
Raskind MA, Peskind ER, Wessel T, and Yuan W (2000) Galantamine in AD: A6-month randomized, placebo-controlled trail with a 6-month extension. The Galantamine USA-1 Study Group. Neurology 54: 2261-2268.
Raymond LA, Tingley WG, Blackstone KW, and Huganir RL (1994) Glutamate receptor modulation by protein phosphorylation. J Physiol (Lond) 88: 181-192.
Riekkinen P Jr, Ikonen S, and Riekkinen M (1998) D-Cycloserine, a partial NMDA receptor-associated glycine-B site agonist, enhances reversal learning, but a cholinesterase inhibitor and nicotine has no effect. Neuroreport 9: 3647-3651.[Medline]
Samochocki M, Hoffle A, Fehrenbacher A, Jostock R, Ludwig J, Christner C, Radina M, Zerlin M, Ullmer C, Pereira EF, et al. (2003) Galantamine is an allosterically potentiating ligand of neuronal nicotinic but not of muscarinic acetylcholine receptors. J Pharmacol Exp Ther 305: 1024-1036.
Santos MD, Alkondon M, Pereira EF, Aracava Y, Eisenberg HM, Maelicke A, and Albuquerque EX (2002) The nicotinic allosteric potentiating ligand galantamine facilitates synaptic transmission in the mammalian central nervous system. Mol Pharmacol 61: 1222-1234.
Schrattenholz A, Pereira EFR, Roth U, Weber KH, Albuquerque EX, and Maelicke A (1996) Agonist responses of neuronal nicotinic acetylcholine receptors are potentiated by a novel class of allosterically acting ligands. Mol Pharmacol 49: 1-6.[Abstract]
Schwartz BL, Hashtroudi S, Herting RL, Handerson H, and Deutch SI (1991) Glycine prodrug facilitates memory retrieval in humans. Neurology 41: 1341-1343.
Schwartz BL, Hashtroudi S, Herting RL, Schwartz P, and Deutsch SI (1996) d-Cycloserine enhances implicit memory in Alzheimer's patients. Neurology 46: 420-424.
Snell LD, Iorio KR, Tabakoff B, and Hoffman PL (1994) Protein kinase C activation attenuates N-methyl-D-aspartate-induced increases in intracellular calcium in cerebellar granule cells. J Neurochem 62: 1783-1789.[Medline]
Tariot PN, Farlow MR, Grossberg GT, Graham SM, MacDonald S, and Gergel I (2004) Memantine treatment in patients with moderate to severe Alzheimer's disease already receiving donepezil: a randomized controlled trail. J Am Med Assoc 291: 317-324.
Tariot PN, Solomon PR, Morris JC, Kershaw P, Lilienfeld S, and Ding C (2000) A 5-month, randomized, placebo-controlled trail of galantamine in AD. The Galantamine USA-10 Study Group. Neurology 54: 2269-2276.
Thomson T, Kewitz H, Bickel V, Straschill M, and Holl G (1991) Preclinical and clinical studies with galantamine, in Cholinergic Basis of Alzheimer's Disease (Becker R and Giacobini E eds), pp 328-336, Birhauser, Boston.
Urushihara H, Tohda M, and Nomura Y (1992) Selective potentiation of N-methyl-D-aspartate-induced current by protein kinase C in Xenopus oocytes injected with rat brain RNA. J Biol Chem 267: 11697-11700.
Vidal C and Changeux JP (1996) Neuronal nicotinic acetylcholine receptors in the brain. News Physiol Sci 11: 202-208.
Wagner DA and Leonard JP (1996) Effect of protein kinase-C activation on the Mg2+-sensitivity of cloned NMDA receptors. Neuropharmacology 35: 29-36.[CrossRef][Medline]
Wang LY and MacDonald JF (1995) Modulation by magnesium of the affinity of NMDA receptors for glycine in murine hippocampal neurones. J Physiol (Lond) 486: 83-95.
Woodruff-Pak DS and Hinchliffe RM (1997) Mecamylamine- or scopolamine-induced learning impairment: ameliorated by nefiracetam. Psychopharmacology 131: 130-139.[CrossRef][Medline]
Xiong ZG, Raouf R, Lu WY, Wang LY, Orser BA, Dudek EM, Browning MD, and MacDonald JF (1998) Regulation of N-methyl-D-aspartate receptor function by constitutively active protein kinase C. Mol Pharmacol 54: 1055-1063.
Zhao X, Kuryatov A, Lindstorm JM, Yeh JZ, and Narahashi T (2001) Nootropic drug modulation of neuronal nicotinic acetylcholine receptors in rat cortical neurons. Mol Pharmacol 59: 674-683.
This article has been cited by other articles:
|
|
 |
|
  S. Lorrio, M. Sobrado, E. Arias, J. M. Roda, A. G. Garcia, and M. G. Lopez Galantamine Postischemia Provides Neuroprotection and Memory Recovery against Transient Global Cerebral Ischemia in Gerbils J. Pharmacol. Exp. Ther., August 1, 2007; 322(2): 591 - 599. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. M. Broad, R. Zwart, K. H. Pearson, M. Lee, L. Wallace, G. I. McPhie, R. Emkey, S. P. Hollinshead, C. P. Dell, S. R. Baker, et al. Identification and Pharmacological Profile of a New Class of Selective Nicotinic Acetylcholine Receptor Potentiators J. Pharmacol. Exp. Ther., September 1, 2006; 318(3): 1108 - 1117. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Geerts and G. T. Grossberg Pharmacology of Acetylcholinesterase Inhibitors and N-methyl-D-aspartate Receptors for Combination Therapy in the Treatment of Alzheimer's Disease J. Clin. Pharmacol., July 1, 2006; 46(suppl_1): 8S - 16S. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Unger, M. M. Svedberg, W.-F. Yu, M. M. Hedberg, and A. Nordberg Effect of Subchronic Treatment of Memantine, Galantamine, and Nicotine in the Brain of Tg2576 (APPswe) Transgenic Mice J. Pharmacol. Exp. Ther., April 1, 2006; 317(1): 30 - 36. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Moriguchi, X. Zhao, W. Marszalec, J. Z. Yeh, and T. Narahashi Modulation of N-Methyl-D-aspartate Receptors by Donepezil in Rat Cortical Neurons J. Pharmacol. Exp. Ther., October 1, 2005; 315(1): 125 - 135. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
 |
 |
|
 |
 |
 |
