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Glucose attenuates the effect of combined muscarinic-nicotinic receptor blockade on spontaneous alternation
ELSEVIER
ejp European Journal of Pharmacology 256 (1994) 31-36
Glucose attenuates the effect of combined muscarinic-nicotinic receptor blockade on spontaneous alternation Michael E. Ragozzino, Gloria Arankowsky-Sandoval 1, Paul E. Gold * Department of Psychology, Gilmer Hall, University of Virginia, Charlottest~ille, VA 22903, USA (Received 23 December 1993; accepted 28 December 1993)
Abstract
Glucose administration reverses the effects of both muscarinic and nicotinic cholinergic receptor antagonists on memory and other measures. In experiment 1, we found that glucose attenuated impairments on spontaneous alternation after muscarinic (scopolamine, 0.5 mg/kg) or nicotinic (mecamylamine, 5.0 mg/kg) receptor blockade. In experiment 2, we examined whether glucose could reverse the spontaneous alternation impairments produced by combined muscarinic-nicotinic receptor blockade. Scopolamine (0.1 mg/kg) and mecamylamine (2.5 mg/kg)when administered separately did not modify alternation performance, but when coadministered they decreased spontaneous alternation scores. This decrease was attenuated by glucose at 100, 300, 500 and 3000 mg/kg. These findings suggest that glucose may attenuate the behavioral impairment by enhancing cholinergic activity and/or other neurotransmitter systems.
Key words: Glucose; Mecamylamine; Scopolamine; Spontaneous alternation; Memory
I. Introduction
Several lines of evidence indicate that circulating glucose levels regulate memory processes. For example, post-training glucose treatment enhances memory in rodents for both aversive and appetitive tasks (e.g., Gold, 1986; Messier and Destrade, 1988; Packard and White, 1990; Means and Fernandez, 1992). In elderly humans, glucose ingestion shortly before or after the presentation of verbal test material enhances memory assessed 24 h later (Manning et al., 1992; Parsons and Gold, 1992). Furthermore, blood glucose levels after training in rodents and glucose tolerance in elderly humans can predict performance in memory tasks (Hall and Gold, 1986; Hall et al., 1989; Manning et al., 1990; Hall and Gold, 1992). Experiments with rodents indicate there are two effective ranges of glucose treatment. Glucose enhances learning and memory when given in the 100-300
* Corresponding author. Tel. (804) 924-0685; fax (804) 982-4785, [email protected]. 1 Now at Centro de Investigaciones Regionales, Universidad Autonomo del Yucatan, Merida, Mexico. 0014-2999/94/$07.00 © 1994 Elsevier Science B.V. All rights reserved SSDI 0 0 1 4 - 2 9 9 9 ( 9 4 ) 0 0 0 1 7 - 2
mg/kg (Gold, 1986; Packard and White, 1990; White, 1991; Means and Fernandez, 1992) and the 2-3 g/kg range of doses (Messier and White, 1987; Messier and Destrade, 1988; Packard and White, 1990; White, 1991). There is evidence suggesting that the higher doses act indirectly on brain mechanisms involved in learning and memory (White, 1991). Conversely, the lower doses may have direct central actions. On the basis of these findings, several experiments have attempted to identify the brain mechanisms that may underlie glucose enhancement of memory storage processes. One set of results suggests that glucose facilitates mnemonic processing by enhancing central cholinergic activity. For example, glucose attenuates amnesia induced by the muscarinic receptor antagonist, scopolamine (Stone et al., 1988a, 1991; Messier et al., 1990). Similar to its attenuation of scopolamineinduced amnesia, glucose also attenuates spontaneous alternation and inhibitory avoidance impairments produced by the nicotinic receptor antagonist, mecamylamine (Ragozzino and Gold, 1991). The interaction of glucose with cholinergic drugs is not limited to deficits in memory tasks. In addition, glucose attenuates scopolamine-induced hyperactivity and facilitates the onset and severity of tremors caused by physostigmine, an
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indirect acetylcholine receptor agonist (Stone et al., 1987, 1988b). Furthermore, glucose treatment, at doses which enhance memory, modifies acetylcholine synthesis and potentiates the scopolamine-induced increase in acetylcholine output (Messier et al., 1990; Durkin et al., 1992). Thus, glucose attenuates the behavioral effects of both a muscarinic and nicotinic acetylcholine receptor antagonist, as well as alters acetylcholine synthesis and output after scopolamine treatment. Riekkinen and colleagues (1990) found that injections of scopolamine and mecamylamine impair learning at doses which individually do not affect learning, in inhibitory avoidance and water-maze tasks. Similarly, combined administration of subthreshold doses of scopolamine and mecamylamine also impair performance in a radial-arm maze task (Levin et al., 1989). These findings indicate that subthreshold doses of muscarinic and nicotinic receptor antagonists administered concurrently have additive effects. In the present study, we examined whether subthreshold doses of scopolamine and mecamylamine would impair spontaneous alternation behavior and whether the impairment could be attenuated by glucose treatment. One possible explanation for glucose attenuation of the behavioral effects of muscarinic and nicotinic receptor antagonists is that, when one class of cholinergic receptors is blocked, glucose treatment results in an enhancement of cholinergic activity which acts on the other receptor class, leading to an attenuation of the behavioral impairment. For example, administering scopolamine and glucose would lead to blockade of muscarinic receptors and at the same time enhance cholinergic activity to activate the nicotinic receptors. Administering mecamylamine and glucose would lead to blockade of nicotinic receptors and increase cholinergic activity to activate muscarinic receptors. Our hypothesis was that glucose treatment would not attenuate a behavioral impairment induced by combined treatment with muscarinic and nicotinic receptor antagonists. In experiment 1, we replicated the glucose attenuation of scopolamine- and mecamylamine-induced impairments of spontaneous alternation performance. In experiment 2, we found that glucose also reversed spontaneous alternation reductions caused by combined scopolamine and mecamylamine treatment, thus contradicting the above hypothesis.
2. Materials and methods 2.1. Subjects
Six-week-old male mice (ICR) were obtained from Hilltop Laboratories (Scottsdale, PA). The mice were housed in groups of four and maintained on a 12:12 h
light-dark cycle (lights on 07.00 h), with free access to food and water. All behavioral testing occurred between 08.00 and 13.00 h. 2.2. Drugs
All drugs were dissolved in sterile saline and administered subcutaneously. Scopolamine hydrochloride and mecamylamine hydrochloride were obtained from the Sigma Chemical Company and D-glucose from the J.T. Baker Chemical Company. 2.3. Spontaneous alternation testing
Each mouse received two concomitant injections 30 min before behavioral testing. One injection was administered in a volume of 0.1 m l / k g and the other injection was administered in a volume of 0.5 ml/kg. In experiment 1, glucose effects in spontaneous alternation following mecamylamine and scopolamine treatment was examined. Each mouse received one of the following treatments: saline-saline, glucose (100 r a g / kg)-saline, mecamylamine (5 mg/kg)-saline, scopolamine (0.5 mg/kg)-saline, mecamylamine (5 m g / k g ) glucose (100 m g / k g ) or scopolamine (0.5 m g / k g ) glucose (100 mg/kg). In experiment 2, glucose effects in spontaneous alternation following combined scopolamine and mecamylamine treatment were examined. In the control groups, each mouse received an injection of saline and an injection of one of the following: saline, scopolamine (0.1 mg/kg), mecamylamine (2.5 mg/kg), scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg), glucose (100 mg/kg), glucose (200 mg/kg), glucose (300 mg/kg), glucose (500 mg/kg), glucose (2 g/kg) or glucose (3 g/kg). In the experimental groups, each mouse received an injection of scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg) plus an injection of glucose 100, 200, 300, 500, 2000 or 3000 mg/kg. Mice coadministered scopolamine and mecamylamine received them as a single injection. In both experiments, some mice within a drug group, selected at random, received a treatment in the larger volume and others received the same treatment in the smaller volume. No behavioral differences were observed between any one treatment given in the larger volume and the same treatment in the smaller volume. The number of mice in a group ranged from 7 to 13. Spontaneous alternation performance was observed in a Y-maze using procedures based on those of Sarter et al. (1988). The Y-maze consisted of 3 trough-shaped arms. The length of each arm was 60 cm, the height of the maze was 17.5 cm, the width of the floor was 3.5 cm, and the width of the ceiling was 14 cm. The arms converged on a roughly triangular central area that was 4 cm along its longest axis. The ceiling was composed
M.E. Ragozzino et al. / European Journal of Pharmacology 256 (1994) 31-36
of translucent, dark acrylic. The mice were placed in one arm and allowed to traverse the maze freely for 8 min while the number and the sequence of entries were recorded. An alternation was defined as the consecutive entry into all 3 arms on overlapping triplet sets. Three consecutive arm choices within the total set of arm choices made up a triplet set. With this procedure, possible alternation sequences are equal to the number of arms entered minus 2, and the percentage of alternation behavior is equal to the ratio of (actual alternations/possible alternations) × 100. For example, if an animal in a session chose the following sequence of arms: ABACBACABC, then the animal would have alternated 6 out of 8 times, with a score of 75%.
2.4. Statistical analysis The percent alternation scores were analyzed using a one-way analysis of variance, followed by protected t-tests (Bruning and Kintz, 1987). The number of arm entries was analyzed by two-tailed t-tests.
3. Results
As illustrated in Fig. 1, both the muscarinic receptor antagonist, scopolamine, and the nicotinic receptor antagonist, mecamylamine, reduced spontaneous alternation performance. These effects were reversed by concomitant administration of glucose. Analysis of the percent alternation scores revealed a significant group effect (F(5,52) = 9.80, P < 0.0001). The saline controls
Z
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SAL GLU MEG MEC SCP SCP SAL SAL SAL GLU SAL GLU
Fig. 1. Glucose reversal of mecamylamine- and scopolamine-induced spontaneous alternation reductions. Mecamylamine (5.0 mg/kg), as well as scopolamine (0.5 m g / k g ) treatment, significantly reduced spontaneous alternation scores. Systemic glucose (100 m g / k g ) alone did not modify spontaneous alternation performance, but when coadministered with mecamylamine or scopolamine, it significantly increased alternation behavior compared to mecamylamine and scopolamine treatment alone and the score was not significantly different from that of saline controls. Key to abbreviations: SAL = saline; G L U = glucose; MEC = mecamylamine; SCP = scopolamine; o p < 0.05 vs. SAL-SAL; [] P < 0.05 vs. MEC-SAL; o p < 0.05 vs. SCP-SAL.
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had a mean alternation score of 64.2% which was slightly lower than that of controls from previous experiments (Ragozzino and Gold, 1991; Stone et al., 1991). Scopolamine-treated mice alternated significantly less than saline controls ( P < 0.05). Similarly, mecamylamine-treated mice exhibited spontaneous alternation scores significantly lower than those of saline controls ( P < 0.05). Glucose itself did not modify alternation performance as compared to that of the saline controls ( P > 0.05). However, mice receiving simultaneous injections of mecamylamine and glucose had alternation scores significantly higher than those of mecamylamine-saline mice ( P <0.05). Mice injected concomitantly with scopolamine and glucose had alternation scores significantly higher than those of scopolamine-saline mice ( P < 0.05). Analysis of arm entries revealed that all drug groups made fewer arm choices than did the saline controls (mean = 44.3 + 2.5 S.E.M.). The glucose controls (31.7 + 1.5) made significantly fewer choices than did the saline controls ( t ( 1 6 ) = 4.52, P < 0.0003). The number of arms entered by mecamylamine-treated mice (19.9 _+ 2.4) and scopolamine-treated mice (23.0 +_ 3.1) was significantly reduced as compared to those entered by saline-treated mice ( t ( 1 5 ) = 7.03 and t(17)= 5.04, respectively, P < 0.0001 in all cases). Similarly, combined treatment with mecamylamine and glucose (23.5 _+ 1.4) and scopolamine and glucose (25.2 +_ 1.9) significantly lowered the number of arms entered compared to the effect of saline treatment (t(14) = 7.26 and t(18) = 6.18, respectively, P < 0.0001 in all cases). Glucose coadministered with mecamylamine or scopolamine did not significantly modify the number of arm choices compared to mecamylamine or scopolamine with saline treatment ( P > 0.05 in both cases). As in previous experiments (Ragozzino and Gold, 1991), the drugs altered the number of arms entered but this measure did not predict alternation performance. The results from experiment 2 are shown in Figs. 2 and 3. The administration of scopolamine (0.1 m g / k g ) or mecamylamine (2.5 m g / k g ) separately did not alter spontaneous alternation performance. However, the coadministration of scopolamine and mecamylamine at subthreshold doses did impair alternation behavior. This impairment was attenuated by glucose 100 m g / k g and reversed by glucose 300 m g / k g and 500 mg/kg. Similarly, glucose 3000 m g / k g attenuated the impairment caused by the combined treatment with scopolamine and mecamylamine. Glucose 200 m g / k g and 2000 m g / k g did not attenuate the reduced alternation scores seen after combined scopolamine-mecamylamine treatment. None of the glucose doses when they were combined with saline changed spontaneous alternation behavior (see Fig. 3). A one-way analysis of variance for the percent alternation scores indicated a significant group effect
M.E. Ragozzino et al. / European Journal of Pharmacology 256 (1994) 31-36
34
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SAL MEG SCP M-S M-S M-S M-S M-S M-S M-S SAL SAL SAL SAL [100 200 300 500 2000 3000] GLUCOSE (mg/kg)
Fig. 2. Glucose reversal of spontaneous alternation impairments due to coadministration of mecamylamine and scopolamine. Mecamylamine (2.5 m g / k g ) and scopolamine (0.1 m g / k g ) when administered separately did not significantly change alternation performance, but when coadministered significantly reduced alternation performance. Of the glucose doses administered in combination with mecamylamine-scopolamine, 100, 300, 500 m g / k g and 3 g / k g significantly increased alternation scores compared to mecamylamine-scopolamine controls. Only the 300- and 5 0 0 - m g / k g doses had effects not significantly different from those in saline controls. Key to abbreviations: SAL = saline; M E C = mecamylamine; SCP = scopolamine; M-S = mecamylamine-scopolamine; a p < 0.05 vs. S A L / S A L ; o p < 0.05 vs. M - S / S A L .
(F(15,147) = 3.90, P < 0.0001). Saline controls alternated with a score near 70%, as seen in previous experiments (Ragozzino and Gold, 199l; Stone et al., 1991). Injections of scopolamine (0.1 mg/kg)-saline did not change the alternation scores compared to those of the saline controls ( P > 0 . 0 5 ) . Similar to scopolamine-treated mice, animals treated with mecamylamine (2.5 m g / kg)-saline did not have alternation scores significantly different from those of saline controls ( P > 0.05). In
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contrast, combined scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg) treatment significantly reduced spontaneous alternation performance compared to the effect of saline-saline treatment ( P < 0 . 0 5 ) . Mice treated with scopolamine-mecamylamine and glucose 100 mg/kg or 3000 mg/kg had significantly higher alternation scores than did mice treated with saline and scopolamine-mecamylamine, ( P < 0 . 0 5 in both cases), though their scores did not reach the level of the saline controls (P < 0.05 in both cases). Glucose 300 and 500 mg/kg significantly increased alternation scores in mice administered scopolamine-mecamylamine (P < 0.05 in both cases) to values not significantly different from those of the saline controls (P > 0.05 in both cases). Mice injected with scopolaminemecamylamine and glucose 200 mg/kg or 2000 mg/kg had alternation scores not significantly different from those of mice administered scopolamine-mecamylamine (P > 0.05 in both cases). None of the glucose control groups had alternation scores significantly different from those of saline controls (P > 0.05 in both cases). Comparable to experiment 1, mecamylamine (mean = 27.6 _+ 2.2 S.E.M.) and scopolamine treatment (22.8 _+ 2.5) significantly reduced the number of arm choices compared to those after saline (35.6 _+ 2.0, t(19) = 2.70 and t(17)= 4.04, respectively, P < 0.02 in all cases). Mice receiving combined mecamylamine-scopolamine treatment (22.1 + 2.9) made significantly fewer arm choices than saline-treated mice (t(18)= 3.94, P < 0.001). Glucose coadministered with mecamylamine and scopolamine reduced the number of arms entered at all glucose doses, except 200 and 2000 mg/kg, compared to saline controls (P < 0.05 in all cases). The number of arm choices made by glucose controls, except at the 500 mg/kg dose, was not significantly different from those of saline controls (P > 0.05 in all cases). As in experiment 1, drug effects on the number of arms entered did not predict alternation scores.
40
SAL GLU GLU GLU GLU GLU GLU SAL SAL SAL SAL SAL SAL SAL 100 200 300 500 2000 3000
Fig. 3. Glucose administration alone did not change spontaneous alternation performance. Glucose doses ranging from 100 m g / k g to 3 g / k g did not significantly alter spontaneous alternation performance compared to saline-saline treatment. Key to abbreviations: SAL = saline; G L U = glucose.
The findings from the present experiment demonstrating scopolamine- and mecamylamine-induced deficits in spontaneous alternation performance are similar to previous results indicating that these drugs impair performance in memory tasks (Goldberg et al., 1971; Flood et al., 1981; Levin et al., 1987, 1989; Berz et al., 1992; Decker and Majchrzak, 1992). Coadministration of subthreshold doses of scopolamine and mecamylamine also impairs spontaneous alternation performance. This finding is consistent with a report by Reikkinen and colleagues (1990) that combined mecamylamine-scopolamine treatment disrupts performance in the Morris water maze and inhibitory avoidance tasks. These results suggest that activation of mus-
M.E. Ragozzino et at,./European Journal of Pharmacology 256 (1994) 31-36
carinic and nicotinic receptors contributes to mnemonic processing. Similar to previous observations (Ragozzino and Gold, 1991; Stone et al., 1991), glucose (100 mg/kg) reversed the impairment in spontaneous alternation performance after either scopolamine or mecamylamine administration. In experiment 2, 100 mg/kg of glucose partially reversed the spontaneous alternation impairment due to coadministration of mecamylamine and scopolamine, while higher doses of glucose (300 and 500 mg/kg) completely reversed the spontaneous alternation impairment. Although the 2000 mg/kg dose did not attenuate spontaneous alternation reductions, the 3000 mg/kg dose of glucose, previously found to attenuate scopolamine-induced amnesia (Messier et al., 1990), also partially reversed the spontaneous alternation impairment. White (1991) found that in a conditioned avoidance task, glucose enhances memory at 100, 2000 and 3000 mg/kg. Examining the effects of glucose on cholinergic blockade in a spontaneous alternation task, we observed that glucose was effective at doses similar to those found effective in the conditioned avoidance task. Previous work suggests that the 2000-3000 mg/kg dose of glucose acts peripherally while the lower doses of glucose have direct central actions (Messier and White, 1987; White, 1991). More specifically, White (1991) demonstrated that lesions of the celiac ganglion, through which most of the autonomic nerves leaving the liver pass en route to the central nervous system (Jacobowitz, 1965; Kreulen and Szurszewski, 1979), block the memory-enhancing effect of the high doses of glucose. Despite this possible difference in mechanism between the low and high doses of glucose, both attenuated the effect of muscarinic-nicotinic cholinergic blockade. While glucose attenuated the effect of combined mecamylamine-scopolamine treatment, glucose alone did not enhance spontaneous alternation performance. This is the first demonstration that glucose treatment alone does not enhance spontaneous alternation performance across both ranges of effective doses. The inability of glucose to alter spontaneous alternation does not seem specific to the treatment itself because other treatments known to improve memory similarly fail to enhance spontaneous alternation performance (Stone et al., 1991; Walker et al., 1991). The inability of memory-enhancers to increase spontaneous alternation suggests that the 70-80% alternation rate observed is the maximal performance in this task for rodents. Glucose attenuation of cholinergic receptor blockade might suggest that glucose acts through a cholinergic mechanism. However, several drugs that have their main actions on other neurotransmitter systems reverse scopolamine-induced amnesia (Flood and Cherkin, 1986; Sarter et al., 1988; Walker et al., 1991). Thus, glucose, at either low or high doses, might act on or
35
through multiple neurotransmitter systems at once. If glucose enhances central cholinergic activity to attenuate scopolamine- and mecamylamine-induced changes, then the results from experiment 2 suggest that it is unlikely that the effect is due solely to activation of the class of acetylcholine receptors that is not blocked. An alternative possibility is that glucose enhances cholinergic activity, which not only leads to activation of the unbiocked receptor class, in the presence of scopolamine or mecamylamine, but also competes for the binding sites at which the acetylcholine receptor antagonists act. Durkin and colleagues (1992) demonstrated that glucose enhances the increased acetylcholine output caused by scopolamine. Thus, with coadministration of glucose and scopolamine an increased acetylcholine output may compete with scopolamine for binding to the muscarinic receptors. A similar process may occur with glucose and mecamylamine. Other evidence that the effects of glucose may act through a cholinergic mechanism is that both muscarinic and nicotinic receptor antagonists are known to modify acetylcholine synthesis (Dolezal and Tucek, 1982; Messier et al., 1990; Elrod and Buccafusco, 1991). The change in acetylcholine synthesis caused by acetylcholine receptor antagonists is attenuated by glucose; possibly through increased availability of acetyl CoA (Dolezal and Tucek, 1982; Messier et al., 1990). Moreover, acetylcholine synthesis may also be regulated by acetyl CoA under other conditions, i.e. hypoxia and aging (Gibson and Blass, 1976; Gibson and Peterson, 1981). Because brain acetyl CoA is derived from glucose (Tucek and Cheng, 1974), increased blood glucose levels may enhance the availability of acetyl CoA under conditions which alter acetylcholine synthesis. However, the 3000-mg/kg dose of glucose, which appears to act peripherally, may alter acetylcholine activity through different mechanisms than the lower glucose doses. In conclusion, we found that glucose reverses the scopolamine-induced and mecamylamine-induced impairment of spontaneous alternation. Glucose, at both low and high doses, also reverses spontaneous alternation reductions induced by combined scopolamine-mecamylamine treatment. However, glucose treatment alone, across the range of doses, did not modify spontaneous alternation performance. Whether the glucose reversal of the effect of combined muscarinic-nicotinic cholinergic blockade indicates that glucose acts by enhancing cholinergic activity or other neurotransmitter systems is still to be determined.
5. Acknowledgements Supported ( A G 07648), trainee on an recipient of a
by research grants from NSF (BNS-9012239), NIA O N R (N0001489-J-1216). M.E.R. was a predoctoral N I M H training grant (5-T32-MH18411). G.A.S. was a Fogarty post-doctoral fellowship.
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6. References Berz, S., K. Battig and H. Welzl, 1992, The effects of anticholinergic drugs on delayed time discrimination performance in rats, Physiol. Behav. 51, 493. Bruning, J.L. and B.L. Kintz, 1987, Computational Handbook of Statistics (Scott, Foresman and Co., Glenview) p. 116. Decker, M.W. and M.J. Majchrzak, 1992, Effects of systemic and intracerebroventricular administration of mecamylamine, a nicotinic cholinergic antagonist, on spatial memory in rats, Psychopharmacology 107, 530. Dolezal, V. and S. Tucek, 1982, Effects of choline and glucose on atropine-induced alterations of acetylcholine synthesis and content in the caudate nuclei of rats, Brain Res. 240, 285. Durkin, T.P., C. Messier, P. DeBoer and B.H.C. Westerwink, 1992, Raised glucose levels enhance scopolamine-induced acetylcholine overflow from the hippocampus: an in vivo microdialysis study in the rat, Behav. Brain Res. 49, 181. Elrod, K. and J.J. Buccafusco, 1991, Correlation of the amnestic effects of nicotinic antagonists with inhibition of regional brain acetylcholine synthesis in rats, J. Pharmacol. Exp. Ther. 258, 403. Flood, J.F. and A. Cherkin, 1986, Scopolamine effects on memory retention in mice: a model of dementia?, Behav. Neural Biol. 45, 169. Flood, J.F., D.W. Landry and M.E. Jarvik, 1981, Cholinergic receptor interactions and their effects on long-term memory processing, Brain Res. 215, 177. Gibson, G.E. and J.P. Blass, 1976, Impaired synthesis of acetylcholine in brain accompanying mild hypoxia and hypoglycemia, J. Neurochem. 27, 37. Gibson, G.E. and C. Peterson, 1981, Aging decreases oxidative metabolism and the release and synthesis of acetylcholine, J. Neurochem. 37, 978. Gold, P.E., 1986, Glucose modulation of memory storage processing, Behav. Neural Biol. 45, 342. Goldberg, M.E., K. Sledge, M. Hefner and R.C. Robichaud, 1971, Learning impairment after three classes of agents which modify cholinergic function, Arch. Int. Pharmacodyn. 193, 226. Hall, J.L. and P.E. Gold, 1986, The effects of training, epinephrine, and glucose injections on plasma glucose levels in rats, Behav. Neural Biol. 46, 156. Hall, J.L. and P.E. Gold, 1992, Plasma glucose levels predict the susceptibility of memory enhancement to disruption by adrenergic antagonists, Eur. J. Pharmacol. 221, 365. Hall, J.L., L.A. Gonder-Frederick, W.W. Chewning, J. Silveira and P.E. Gold, 1989, Glucose enhancement of performance on memory tests in young and aged humans, Neuropsychologia 27, 1129. Jacobowitz, D.M., 1965, Histochemical studies of the autonomic innervation of the gut, J. Pharmacol. Exp. Ther. 149, 358. Kreulen, D.L. and J.H. Szurszewski, 1979, Nerve pathways in celiac plexus of the guinea pig, Am. J. Physiol. 237, E90. Levin, E.D., M. Castonguay and G.D. Ellison, 1987, Effects of the nicotinic receptor blocker mecamylamine on radial-arm maze performance in rats, Brain Res. 215, 177. Levin, E.D., S.R. McGurk, D. South and L.L. Butcher, 1989, Effects of combined muscarinic and nicotinic blockade on choice accuracy in the radial-arm maze, Behav. Neural Biol. 51,270.
Manning, C.A., J.L. Hall and P.E. Gold, 1990, Glucose effects on memory and other neuropsychological tests in elderly humans, Psychol. Sci. 5, 307. Manning, C.A., M.W. Parsons and P.E. Gold, 1992, Anterogradc and retrograde enhancement of 24-hour memory by glucose in elderly humans, Behav. Neural Biol. 58, 125. Means, L.W. and T.J. Fernandez, 1992, Daily glucose injections facilitate performance of a win-stay water-escape working memory task in mice, Behav. Neurosci. 106, 345. Messier, C. and C. Destrade, 1988, Improvement of memory for an operant response by post-training glucose in mice, Beh. Brain Res. 3l, 185. Messier, C. and N.M. White, 1987, Memory improvement by glucose, fructose, and two glucose analogs: a possible effect on peripheral glucose transport, Behav. Neural Biol. 48, 104. Messier, C., T. Durkin, O. Mrabet and C. Destrade, 1990, Memoryimproving action of glucose: indirect evidence for a facilitation of hippocampal acetylcholine synthesis, Beh. Brain Res. 39, 135. Packard, M.G. and N.M. White, 1990, Effect of posttraining injections of glucose on acquisition of two appetitive learning tasks, Psychobiology 18, 282. Parsons, M. and P.E. Gold, 1992, Glucose enhancement of memory in elderly humans: An inverted-U dose-response curve, Neurobiol. Aging 13, 401. Ragozzino, M.E. and P.E. Gold, 1991, Glucose effects on mecamylamine-induced memory deficits and decreases in locomotor activity in mice, Behav. Neural Biol. 56, 271. Riekkinen, P., J. Sirvio, M. Aaltonen and P. Riekkinen, 1990, Effects of concurrent manipulations of nicotinic and muscarinic receptors on spatial and passive avoidance learning, Pharmacol. Biochem. Behav. 37, 405. Sarter, M., G. Bodewitz and D.N. Stephens, 1988, Attenuation of scopolamine-induced impairment of spontaneous alternation behaviour by antagonist but not inverse agonist and agonist /3carbolines, Psychopharmacology 94, 491. Stone, W.S., K.L. Cottrill and P.E. Gold, 1987, Glucose and epinephrine attenuation of scopolamine-induced increases in locomotor activity in mice, Neurosci. Res. Commun. 1, 105. Stone, W.S., C.E. Croul and P.E. Gold, 1988a, Attenuation of scopolamine-induced amnesia, Psychopharmacology 96, 417. Stone, W.S., K.L. Cottrill, D.L. Walker and P.E. Gold, 1988b, Blood glucose and brain function: interactions with CNS cholinergic systems, Behav. Neural Biol. 50, 325. Stone, W.S., B. Walser, S.D. Gold and P.E. Gold, 1991, Scopolamine- and morphine-induced impairments of spontaneous alternation performance in mice: reversal with glucose and with cholinergic and adrenergic agonists, Behav. Neurosci. 105, 264. Tucek, S. and S.C. Cheng, 1974, Provenance of the acetyl group of acetylcholine and compartmentation of acetyl-CoA and Krebs cycle intermediates in the brain in vivo, J. Neurochem. 22, 893. Walker, D.L., T. McGlynn, C. Grey, M. Ragozzino and P.E. Gold, 1991, Naloxone modulates the behavioral effects of cholinergic agonists and antagonists, Psychopharmacology 105, 57. White, N.M., 1991, Peripheral and central memory-enhancing actions of glucose, in: Peripheral Signaling of the Brain: Role in Neural-lmmune Interactions and Learning and Memory, eds. R.C.A. Frederickson, J.L. McGaugh and D.L. Fellen (Hogrefe and Huber Publishers, Toronto) p. 421.
ejp European Journal of Pharmacology 256 (1994) 31-36
Glucose attenuates the effect of combined muscarinic-nicotinic receptor blockade on spontaneous alternation Michael E. Ragozzino, Gloria Arankowsky-Sandoval 1, Paul E. Gold * Department of Psychology, Gilmer Hall, University of Virginia, Charlottest~ille, VA 22903, USA (Received 23 December 1993; accepted 28 December 1993)
Abstract
Glucose administration reverses the effects of both muscarinic and nicotinic cholinergic receptor antagonists on memory and other measures. In experiment 1, we found that glucose attenuated impairments on spontaneous alternation after muscarinic (scopolamine, 0.5 mg/kg) or nicotinic (mecamylamine, 5.0 mg/kg) receptor blockade. In experiment 2, we examined whether glucose could reverse the spontaneous alternation impairments produced by combined muscarinic-nicotinic receptor blockade. Scopolamine (0.1 mg/kg) and mecamylamine (2.5 mg/kg)when administered separately did not modify alternation performance, but when coadministered they decreased spontaneous alternation scores. This decrease was attenuated by glucose at 100, 300, 500 and 3000 mg/kg. These findings suggest that glucose may attenuate the behavioral impairment by enhancing cholinergic activity and/or other neurotransmitter systems.
Key words: Glucose; Mecamylamine; Scopolamine; Spontaneous alternation; Memory
I. Introduction
Several lines of evidence indicate that circulating glucose levels regulate memory processes. For example, post-training glucose treatment enhances memory in rodents for both aversive and appetitive tasks (e.g., Gold, 1986; Messier and Destrade, 1988; Packard and White, 1990; Means and Fernandez, 1992). In elderly humans, glucose ingestion shortly before or after the presentation of verbal test material enhances memory assessed 24 h later (Manning et al., 1992; Parsons and Gold, 1992). Furthermore, blood glucose levels after training in rodents and glucose tolerance in elderly humans can predict performance in memory tasks (Hall and Gold, 1986; Hall et al., 1989; Manning et al., 1990; Hall and Gold, 1992). Experiments with rodents indicate there are two effective ranges of glucose treatment. Glucose enhances learning and memory when given in the 100-300
* Corresponding author. Tel. (804) 924-0685; fax (804) 982-4785, [email protected]. 1 Now at Centro de Investigaciones Regionales, Universidad Autonomo del Yucatan, Merida, Mexico. 0014-2999/94/$07.00 © 1994 Elsevier Science B.V. All rights reserved SSDI 0 0 1 4 - 2 9 9 9 ( 9 4 ) 0 0 0 1 7 - 2
mg/kg (Gold, 1986; Packard and White, 1990; White, 1991; Means and Fernandez, 1992) and the 2-3 g/kg range of doses (Messier and White, 1987; Messier and Destrade, 1988; Packard and White, 1990; White, 1991). There is evidence suggesting that the higher doses act indirectly on brain mechanisms involved in learning and memory (White, 1991). Conversely, the lower doses may have direct central actions. On the basis of these findings, several experiments have attempted to identify the brain mechanisms that may underlie glucose enhancement of memory storage processes. One set of results suggests that glucose facilitates mnemonic processing by enhancing central cholinergic activity. For example, glucose attenuates amnesia induced by the muscarinic receptor antagonist, scopolamine (Stone et al., 1988a, 1991; Messier et al., 1990). Similar to its attenuation of scopolamineinduced amnesia, glucose also attenuates spontaneous alternation and inhibitory avoidance impairments produced by the nicotinic receptor antagonist, mecamylamine (Ragozzino and Gold, 1991). The interaction of glucose with cholinergic drugs is not limited to deficits in memory tasks. In addition, glucose attenuates scopolamine-induced hyperactivity and facilitates the onset and severity of tremors caused by physostigmine, an
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indirect acetylcholine receptor agonist (Stone et al., 1987, 1988b). Furthermore, glucose treatment, at doses which enhance memory, modifies acetylcholine synthesis and potentiates the scopolamine-induced increase in acetylcholine output (Messier et al., 1990; Durkin et al., 1992). Thus, glucose attenuates the behavioral effects of both a muscarinic and nicotinic acetylcholine receptor antagonist, as well as alters acetylcholine synthesis and output after scopolamine treatment. Riekkinen and colleagues (1990) found that injections of scopolamine and mecamylamine impair learning at doses which individually do not affect learning, in inhibitory avoidance and water-maze tasks. Similarly, combined administration of subthreshold doses of scopolamine and mecamylamine also impair performance in a radial-arm maze task (Levin et al., 1989). These findings indicate that subthreshold doses of muscarinic and nicotinic receptor antagonists administered concurrently have additive effects. In the present study, we examined whether subthreshold doses of scopolamine and mecamylamine would impair spontaneous alternation behavior and whether the impairment could be attenuated by glucose treatment. One possible explanation for glucose attenuation of the behavioral effects of muscarinic and nicotinic receptor antagonists is that, when one class of cholinergic receptors is blocked, glucose treatment results in an enhancement of cholinergic activity which acts on the other receptor class, leading to an attenuation of the behavioral impairment. For example, administering scopolamine and glucose would lead to blockade of muscarinic receptors and at the same time enhance cholinergic activity to activate the nicotinic receptors. Administering mecamylamine and glucose would lead to blockade of nicotinic receptors and increase cholinergic activity to activate muscarinic receptors. Our hypothesis was that glucose treatment would not attenuate a behavioral impairment induced by combined treatment with muscarinic and nicotinic receptor antagonists. In experiment 1, we replicated the glucose attenuation of scopolamine- and mecamylamine-induced impairments of spontaneous alternation performance. In experiment 2, we found that glucose also reversed spontaneous alternation reductions caused by combined scopolamine and mecamylamine treatment, thus contradicting the above hypothesis.
2. Materials and methods 2.1. Subjects
Six-week-old male mice (ICR) were obtained from Hilltop Laboratories (Scottsdale, PA). The mice were housed in groups of four and maintained on a 12:12 h
light-dark cycle (lights on 07.00 h), with free access to food and water. All behavioral testing occurred between 08.00 and 13.00 h. 2.2. Drugs
All drugs were dissolved in sterile saline and administered subcutaneously. Scopolamine hydrochloride and mecamylamine hydrochloride were obtained from the Sigma Chemical Company and D-glucose from the J.T. Baker Chemical Company. 2.3. Spontaneous alternation testing
Each mouse received two concomitant injections 30 min before behavioral testing. One injection was administered in a volume of 0.1 m l / k g and the other injection was administered in a volume of 0.5 ml/kg. In experiment 1, glucose effects in spontaneous alternation following mecamylamine and scopolamine treatment was examined. Each mouse received one of the following treatments: saline-saline, glucose (100 r a g / kg)-saline, mecamylamine (5 mg/kg)-saline, scopolamine (0.5 mg/kg)-saline, mecamylamine (5 m g / k g ) glucose (100 m g / k g ) or scopolamine (0.5 m g / k g ) glucose (100 mg/kg). In experiment 2, glucose effects in spontaneous alternation following combined scopolamine and mecamylamine treatment were examined. In the control groups, each mouse received an injection of saline and an injection of one of the following: saline, scopolamine (0.1 mg/kg), mecamylamine (2.5 mg/kg), scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg), glucose (100 mg/kg), glucose (200 mg/kg), glucose (300 mg/kg), glucose (500 mg/kg), glucose (2 g/kg) or glucose (3 g/kg). In the experimental groups, each mouse received an injection of scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg) plus an injection of glucose 100, 200, 300, 500, 2000 or 3000 mg/kg. Mice coadministered scopolamine and mecamylamine received them as a single injection. In both experiments, some mice within a drug group, selected at random, received a treatment in the larger volume and others received the same treatment in the smaller volume. No behavioral differences were observed between any one treatment given in the larger volume and the same treatment in the smaller volume. The number of mice in a group ranged from 7 to 13. Spontaneous alternation performance was observed in a Y-maze using procedures based on those of Sarter et al. (1988). The Y-maze consisted of 3 trough-shaped arms. The length of each arm was 60 cm, the height of the maze was 17.5 cm, the width of the floor was 3.5 cm, and the width of the ceiling was 14 cm. The arms converged on a roughly triangular central area that was 4 cm along its longest axis. The ceiling was composed
M.E. Ragozzino et al. / European Journal of Pharmacology 256 (1994) 31-36
of translucent, dark acrylic. The mice were placed in one arm and allowed to traverse the maze freely for 8 min while the number and the sequence of entries were recorded. An alternation was defined as the consecutive entry into all 3 arms on overlapping triplet sets. Three consecutive arm choices within the total set of arm choices made up a triplet set. With this procedure, possible alternation sequences are equal to the number of arms entered minus 2, and the percentage of alternation behavior is equal to the ratio of (actual alternations/possible alternations) × 100. For example, if an animal in a session chose the following sequence of arms: ABACBACABC, then the animal would have alternated 6 out of 8 times, with a score of 75%.
2.4. Statistical analysis The percent alternation scores were analyzed using a one-way analysis of variance, followed by protected t-tests (Bruning and Kintz, 1987). The number of arm entries was analyzed by two-tailed t-tests.
3. Results
As illustrated in Fig. 1, both the muscarinic receptor antagonist, scopolamine, and the nicotinic receptor antagonist, mecamylamine, reduced spontaneous alternation performance. These effects were reversed by concomitant administration of glucose. Analysis of the percent alternation scores revealed a significant group effect (F(5,52) = 9.80, P < 0.0001). The saline controls
Z
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,,=,~ Z
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SAL GLU MEG MEC SCP SCP SAL SAL SAL GLU SAL GLU
Fig. 1. Glucose reversal of mecamylamine- and scopolamine-induced spontaneous alternation reductions. Mecamylamine (5.0 mg/kg), as well as scopolamine (0.5 m g / k g ) treatment, significantly reduced spontaneous alternation scores. Systemic glucose (100 m g / k g ) alone did not modify spontaneous alternation performance, but when coadministered with mecamylamine or scopolamine, it significantly increased alternation behavior compared to mecamylamine and scopolamine treatment alone and the score was not significantly different from that of saline controls. Key to abbreviations: SAL = saline; G L U = glucose; MEC = mecamylamine; SCP = scopolamine; o p < 0.05 vs. SAL-SAL; [] P < 0.05 vs. MEC-SAL; o p < 0.05 vs. SCP-SAL.
33
had a mean alternation score of 64.2% which was slightly lower than that of controls from previous experiments (Ragozzino and Gold, 1991; Stone et al., 1991). Scopolamine-treated mice alternated significantly less than saline controls ( P < 0.05). Similarly, mecamylamine-treated mice exhibited spontaneous alternation scores significantly lower than those of saline controls ( P < 0.05). Glucose itself did not modify alternation performance as compared to that of the saline controls ( P > 0.05). However, mice receiving simultaneous injections of mecamylamine and glucose had alternation scores significantly higher than those of mecamylamine-saline mice ( P <0.05). Mice injected concomitantly with scopolamine and glucose had alternation scores significantly higher than those of scopolamine-saline mice ( P < 0.05). Analysis of arm entries revealed that all drug groups made fewer arm choices than did the saline controls (mean = 44.3 + 2.5 S.E.M.). The glucose controls (31.7 + 1.5) made significantly fewer choices than did the saline controls ( t ( 1 6 ) = 4.52, P < 0.0003). The number of arms entered by mecamylamine-treated mice (19.9 _+ 2.4) and scopolamine-treated mice (23.0 +_ 3.1) was significantly reduced as compared to those entered by saline-treated mice ( t ( 1 5 ) = 7.03 and t(17)= 5.04, respectively, P < 0.0001 in all cases). Similarly, combined treatment with mecamylamine and glucose (23.5 _+ 1.4) and scopolamine and glucose (25.2 +_ 1.9) significantly lowered the number of arms entered compared to the effect of saline treatment (t(14) = 7.26 and t(18) = 6.18, respectively, P < 0.0001 in all cases). Glucose coadministered with mecamylamine or scopolamine did not significantly modify the number of arm choices compared to mecamylamine or scopolamine with saline treatment ( P > 0.05 in both cases). As in previous experiments (Ragozzino and Gold, 1991), the drugs altered the number of arms entered but this measure did not predict alternation performance. The results from experiment 2 are shown in Figs. 2 and 3. The administration of scopolamine (0.1 m g / k g ) or mecamylamine (2.5 m g / k g ) separately did not alter spontaneous alternation performance. However, the coadministration of scopolamine and mecamylamine at subthreshold doses did impair alternation behavior. This impairment was attenuated by glucose 100 m g / k g and reversed by glucose 300 m g / k g and 500 mg/kg. Similarly, glucose 3000 m g / k g attenuated the impairment caused by the combined treatment with scopolamine and mecamylamine. Glucose 200 m g / k g and 2000 m g / k g did not attenuate the reduced alternation scores seen after combined scopolamine-mecamylamine treatment. None of the glucose doses when they were combined with saline changed spontaneous alternation behavior (see Fig. 3). A one-way analysis of variance for the percent alternation scores indicated a significant group effect
M.E. Ragozzino et al. / European Journal of Pharmacology 256 (1994) 31-36
34
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SAL MEG SCP M-S M-S M-S M-S M-S M-S M-S SAL SAL SAL SAL [100 200 300 500 2000 3000] GLUCOSE (mg/kg)
Fig. 2. Glucose reversal of spontaneous alternation impairments due to coadministration of mecamylamine and scopolamine. Mecamylamine (2.5 m g / k g ) and scopolamine (0.1 m g / k g ) when administered separately did not significantly change alternation performance, but when coadministered significantly reduced alternation performance. Of the glucose doses administered in combination with mecamylamine-scopolamine, 100, 300, 500 m g / k g and 3 g / k g significantly increased alternation scores compared to mecamylamine-scopolamine controls. Only the 300- and 5 0 0 - m g / k g doses had effects not significantly different from those in saline controls. Key to abbreviations: SAL = saline; M E C = mecamylamine; SCP = scopolamine; M-S = mecamylamine-scopolamine; a p < 0.05 vs. S A L / S A L ; o p < 0.05 vs. M - S / S A L .
(F(15,147) = 3.90, P < 0.0001). Saline controls alternated with a score near 70%, as seen in previous experiments (Ragozzino and Gold, 199l; Stone et al., 1991). Injections of scopolamine (0.1 mg/kg)-saline did not change the alternation scores compared to those of the saline controls ( P > 0 . 0 5 ) . Similar to scopolamine-treated mice, animals treated with mecamylamine (2.5 m g / kg)-saline did not have alternation scores significantly different from those of saline controls ( P > 0.05). In
z
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contrast, combined scopolamine (0.1 mg/kg)-mecamylamine (2.5 mg/kg) treatment significantly reduced spontaneous alternation performance compared to the effect of saline-saline treatment ( P < 0 . 0 5 ) . Mice treated with scopolamine-mecamylamine and glucose 100 mg/kg or 3000 mg/kg had significantly higher alternation scores than did mice treated with saline and scopolamine-mecamylamine, ( P < 0 . 0 5 in both cases), though their scores did not reach the level of the saline controls (P < 0.05 in both cases). Glucose 300 and 500 mg/kg significantly increased alternation scores in mice administered scopolamine-mecamylamine (P < 0.05 in both cases) to values not significantly different from those of the saline controls (P > 0.05 in both cases). Mice injected with scopolaminemecamylamine and glucose 200 mg/kg or 2000 mg/kg had alternation scores not significantly different from those of mice administered scopolamine-mecamylamine (P > 0.05 in both cases). None of the glucose control groups had alternation scores significantly different from those of saline controls (P > 0.05 in both cases). Comparable to experiment 1, mecamylamine (mean = 27.6 _+ 2.2 S.E.M.) and scopolamine treatment (22.8 _+ 2.5) significantly reduced the number of arm choices compared to those after saline (35.6 _+ 2.0, t(19) = 2.70 and t(17)= 4.04, respectively, P < 0.02 in all cases). Mice receiving combined mecamylamine-scopolamine treatment (22.1 + 2.9) made significantly fewer arm choices than saline-treated mice (t(18)= 3.94, P < 0.001). Glucose coadministered with mecamylamine and scopolamine reduced the number of arms entered at all glucose doses, except 200 and 2000 mg/kg, compared to saline controls (P < 0.05 in all cases). The number of arm choices made by glucose controls, except at the 500 mg/kg dose, was not significantly different from those of saline controls (P > 0.05 in all cases). As in experiment 1, drug effects on the number of arms entered did not predict alternation scores.
40
SAL GLU GLU GLU GLU GLU GLU SAL SAL SAL SAL SAL SAL SAL 100 200 300 500 2000 3000
Fig. 3. Glucose administration alone did not change spontaneous alternation performance. Glucose doses ranging from 100 m g / k g to 3 g / k g did not significantly alter spontaneous alternation performance compared to saline-saline treatment. Key to abbreviations: SAL = saline; G L U = glucose.
The findings from the present experiment demonstrating scopolamine- and mecamylamine-induced deficits in spontaneous alternation performance are similar to previous results indicating that these drugs impair performance in memory tasks (Goldberg et al., 1971; Flood et al., 1981; Levin et al., 1987, 1989; Berz et al., 1992; Decker and Majchrzak, 1992). Coadministration of subthreshold doses of scopolamine and mecamylamine also impairs spontaneous alternation performance. This finding is consistent with a report by Reikkinen and colleagues (1990) that combined mecamylamine-scopolamine treatment disrupts performance in the Morris water maze and inhibitory avoidance tasks. These results suggest that activation of mus-
M.E. Ragozzino et at,./European Journal of Pharmacology 256 (1994) 31-36
carinic and nicotinic receptors contributes to mnemonic processing. Similar to previous observations (Ragozzino and Gold, 1991; Stone et al., 1991), glucose (100 mg/kg) reversed the impairment in spontaneous alternation performance after either scopolamine or mecamylamine administration. In experiment 2, 100 mg/kg of glucose partially reversed the spontaneous alternation impairment due to coadministration of mecamylamine and scopolamine, while higher doses of glucose (300 and 500 mg/kg) completely reversed the spontaneous alternation impairment. Although the 2000 mg/kg dose did not attenuate spontaneous alternation reductions, the 3000 mg/kg dose of glucose, previously found to attenuate scopolamine-induced amnesia (Messier et al., 1990), also partially reversed the spontaneous alternation impairment. White (1991) found that in a conditioned avoidance task, glucose enhances memory at 100, 2000 and 3000 mg/kg. Examining the effects of glucose on cholinergic blockade in a spontaneous alternation task, we observed that glucose was effective at doses similar to those found effective in the conditioned avoidance task. Previous work suggests that the 2000-3000 mg/kg dose of glucose acts peripherally while the lower doses of glucose have direct central actions (Messier and White, 1987; White, 1991). More specifically, White (1991) demonstrated that lesions of the celiac ganglion, through which most of the autonomic nerves leaving the liver pass en route to the central nervous system (Jacobowitz, 1965; Kreulen and Szurszewski, 1979), block the memory-enhancing effect of the high doses of glucose. Despite this possible difference in mechanism between the low and high doses of glucose, both attenuated the effect of muscarinic-nicotinic cholinergic blockade. While glucose attenuated the effect of combined mecamylamine-scopolamine treatment, glucose alone did not enhance spontaneous alternation performance. This is the first demonstration that glucose treatment alone does not enhance spontaneous alternation performance across both ranges of effective doses. The inability of glucose to alter spontaneous alternation does not seem specific to the treatment itself because other treatments known to improve memory similarly fail to enhance spontaneous alternation performance (Stone et al., 1991; Walker et al., 1991). The inability of memory-enhancers to increase spontaneous alternation suggests that the 70-80% alternation rate observed is the maximal performance in this task for rodents. Glucose attenuation of cholinergic receptor blockade might suggest that glucose acts through a cholinergic mechanism. However, several drugs that have their main actions on other neurotransmitter systems reverse scopolamine-induced amnesia (Flood and Cherkin, 1986; Sarter et al., 1988; Walker et al., 1991). Thus, glucose, at either low or high doses, might act on or
35
through multiple neurotransmitter systems at once. If glucose enhances central cholinergic activity to attenuate scopolamine- and mecamylamine-induced changes, then the results from experiment 2 suggest that it is unlikely that the effect is due solely to activation of the class of acetylcholine receptors that is not blocked. An alternative possibility is that glucose enhances cholinergic activity, which not only leads to activation of the unbiocked receptor class, in the presence of scopolamine or mecamylamine, but also competes for the binding sites at which the acetylcholine receptor antagonists act. Durkin and colleagues (1992) demonstrated that glucose enhances the increased acetylcholine output caused by scopolamine. Thus, with coadministration of glucose and scopolamine an increased acetylcholine output may compete with scopolamine for binding to the muscarinic receptors. A similar process may occur with glucose and mecamylamine. Other evidence that the effects of glucose may act through a cholinergic mechanism is that both muscarinic and nicotinic receptor antagonists are known to modify acetylcholine synthesis (Dolezal and Tucek, 1982; Messier et al., 1990; Elrod and Buccafusco, 1991). The change in acetylcholine synthesis caused by acetylcholine receptor antagonists is attenuated by glucose; possibly through increased availability of acetyl CoA (Dolezal and Tucek, 1982; Messier et al., 1990). Moreover, acetylcholine synthesis may also be regulated by acetyl CoA under other conditions, i.e. hypoxia and aging (Gibson and Blass, 1976; Gibson and Peterson, 1981). Because brain acetyl CoA is derived from glucose (Tucek and Cheng, 1974), increased blood glucose levels may enhance the availability of acetyl CoA under conditions which alter acetylcholine synthesis. However, the 3000-mg/kg dose of glucose, which appears to act peripherally, may alter acetylcholine activity through different mechanisms than the lower glucose doses. In conclusion, we found that glucose reverses the scopolamine-induced and mecamylamine-induced impairment of spontaneous alternation. Glucose, at both low and high doses, also reverses spontaneous alternation reductions induced by combined scopolamine-mecamylamine treatment. However, glucose treatment alone, across the range of doses, did not modify spontaneous alternation performance. Whether the glucose reversal of the effect of combined muscarinic-nicotinic cholinergic blockade indicates that glucose acts by enhancing cholinergic activity or other neurotransmitter systems is still to be determined.
5. Acknowledgements Supported ( A G 07648), trainee on an recipient of a
by research grants from NSF (BNS-9012239), NIA O N R (N0001489-J-1216). M.E.R. was a predoctoral N I M H training grant (5-T32-MH18411). G.A.S. was a Fogarty post-doctoral fellowship.
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Manning, C.A., J.L. Hall and P.E. Gold, 1990, Glucose effects on memory and other neuropsychological tests in elderly humans, Psychol. Sci. 5, 307. Manning, C.A., M.W. Parsons and P.E. Gold, 1992, Anterogradc and retrograde enhancement of 24-hour memory by glucose in elderly humans, Behav. Neural Biol. 58, 125. Means, L.W. and T.J. Fernandez, 1992, Daily glucose injections facilitate performance of a win-stay water-escape working memory task in mice, Behav. Neurosci. 106, 345. Messier, C. and C. Destrade, 1988, Improvement of memory for an operant response by post-training glucose in mice, Beh. Brain Res. 3l, 185. Messier, C. and N.M. White, 1987, Memory improvement by glucose, fructose, and two glucose analogs: a possible effect on peripheral glucose transport, Behav. Neural Biol. 48, 104. Messier, C., T. Durkin, O. Mrabet and C. Destrade, 1990, Memoryimproving action of glucose: indirect evidence for a facilitation of hippocampal acetylcholine synthesis, Beh. Brain Res. 39, 135. Packard, M.G. and N.M. White, 1990, Effect of posttraining injections of glucose on acquisition of two appetitive learning tasks, Psychobiology 18, 282. Parsons, M. and P.E. Gold, 1992, Glucose enhancement of memory in elderly humans: An inverted-U dose-response curve, Neurobiol. Aging 13, 401. Ragozzino, M.E. and P.E. Gold, 1991, Glucose effects on mecamylamine-induced memory deficits and decreases in locomotor activity in mice, Behav. Neural Biol. 56, 271. Riekkinen, P., J. Sirvio, M. Aaltonen and P. Riekkinen, 1990, Effects of concurrent manipulations of nicotinic and muscarinic receptors on spatial and passive avoidance learning, Pharmacol. Biochem. Behav. 37, 405. Sarter, M., G. Bodewitz and D.N. Stephens, 1988, Attenuation of scopolamine-induced impairment of spontaneous alternation behaviour by antagonist but not inverse agonist and agonist /3carbolines, Psychopharmacology 94, 491. Stone, W.S., K.L. Cottrill and P.E. Gold, 1987, Glucose and epinephrine attenuation of scopolamine-induced increases in locomotor activity in mice, Neurosci. Res. Commun. 1, 105. Stone, W.S., C.E. Croul and P.E. Gold, 1988a, Attenuation of scopolamine-induced amnesia, Psychopharmacology 96, 417. Stone, W.S., K.L. Cottrill, D.L. Walker and P.E. Gold, 1988b, Blood glucose and brain function: interactions with CNS cholinergic systems, Behav. Neural Biol. 50, 325. Stone, W.S., B. Walser, S.D. Gold and P.E. Gold, 1991, Scopolamine- and morphine-induced impairments of spontaneous alternation performance in mice: reversal with glucose and with cholinergic and adrenergic agonists, Behav. Neurosci. 105, 264. Tucek, S. and S.C. Cheng, 1974, Provenance of the acetyl group of acetylcholine and compartmentation of acetyl-CoA and Krebs cycle intermediates in the brain in vivo, J. Neurochem. 22, 893. Walker, D.L., T. McGlynn, C. Grey, M. Ragozzino and P.E. Gold, 1991, Naloxone modulates the behavioral effects of cholinergic agonists and antagonists, Psychopharmacology 105, 57. White, N.M., 1991, Peripheral and central memory-enhancing actions of glucose, in: Peripheral Signaling of the Brain: Role in Neural-lmmune Interactions and Learning and Memory, eds. R.C.A. Frederickson, J.L. McGaugh and D.L. Fellen (Hogrefe and Huber Publishers, Toronto) p. 421.