大四下毒理文獻分析neuroscience

1、Neuroscience 147 (2007) 853 864CLIOOL AND VITAMIN B12 (COBALAMIN) SYNERGISTICALLY RESCUE THE LEAD-INDUCED IMPAIRMENTS OF SYNAPTIC PLASTICITY IN HIPPOCAMPAL DENTATE GYRUS AREAOF THE ANESTHETIZED RATS IN VIVOW.-H. CHEN, M. WANG, S.-S. YU, L. SU, D.-M. ZHU, J.-Q. SHE, X.-J. CAO AND D.-Y. RUAN*School of

2、 Life Sciences, University of Science and Technology of, Hefei, Anhui, 230027, PR AbstractLead (Pb2 ) exposure in development induces im- pairments of synaptic plasticity in the hippocampal dentate gyrus (DG) area of the anesthetized rats in vivo. The common chelating agents have many adverse effect

3、s and are incapa- ble of alleviating lead-induced neurotoxicity. Recently, CQ, clio ol (5-chloro-7-iodo-8-hydroxy-oline), which is a transition metal ion chelator and/or ionophore with low affin- ity for metal ions, has yielded some promising results in animal ms and clinical trials related to dysfu

4、nctions of metal ions. In addition, CQ-associated side effects are be- lieved to be overcome with vitamin B12 (VB12) supplemen- tation. To determine whether CQ treatment could rescue im- pairments of synaptic plasticity induced by chronic Pb2 exposure, we investigated the input/output functions (I/O

5、s), paired-pulse reactions (PPRs) and long-term potentiation (LTP) of different treatment groups in hippocampal DG area of the anesthetized rat in vivo by recording field potentials and measured hippocampal Pb2 concentrations of different treatment groups by PlasmaQuad 3 inductive coupled plasma mas

6、s spectroscopy. The results show: CQ alone does not rescue the lead-induced impairments of synaptic plasticity in hippocampal DG area of the anesthetized rats in vivo; VB12 alone partly rescues the lead-induced impair- ments of LTP; however the co-administration of CQ and VB12 totally rescues these

7、impairments of synaptic plasticity and moreover, the effects of CQ and VB12 co-administration are specific to the lead-exposed animals. 2007 IBRO. Published by Elsevier . .Key words: lead, cliool, dentate gyrus, hippocampus, rat.Lead (Pb2 ) is one of the most important neurotoxic met- als in the env

8、ironment. It is now well established that the chronic Pb2 exposure in development produces cognitive*Corresponding author. Tel: 86-551-3606374 or 86-551-3601459; fax: 86-551-3601443. address: ruandy (D.-Y. Ruan).Abbreviations: AD, Alzheimers disease; A , -amyloid protein; control CQ VB12, control wi

9、th clio ol and vitamin B12 group; CQ, cliool (5-chloro-7-iodo-8-hydroxy- oline); DG, dentate gy- rus; fEPSP, field excitatory postsynaptic potential; HD, Huntingtons disease; HFS, high frequency stimulus; I/O, input/output function; IPI, interpulse interval; lead CQ, lead-exposed with cliool group;

10、lead CQ VB12, lead-exposed with clio ol and vitamin B12 group; lead VB12, lead-exposed with vitamin B12 group; LTP, long-term potentiation; NMDA, N-methyl-D-aspartate; PD, Parkinsons disease; PPR, paired-pulse reaction; PS, population spike; SMON, subacute myelo-optic neuropathy; VB12, vitamin B12.d

11、eficits and neurobehavioral dysfunction in children and in a lot of animal species (Cory-Slechta, 1990; Bellinger et al., 1991; Banks et al., 1997). Significant cognitive impair- ment of develo brain is reported at blood Pb2 levels as low as 10 g/dl in children (Needleman and Gatsonis, 1990; Belling

12、er et al., 1991) and animal studies have sup- ported these findings, indicating that developmental expo- sure to Pb2 resulting in environmentally relevant blood concentrations also produces cognitive dysfunction (Cory- Slechta et al., 1985, Rice, 1993). In the Pb-exposed ani- mals a blood lead conce

13、ntration of 30 g/dl compares well with the about 10 g/dl blood concentration measured in Pb-exposed children. The hippocampus, which is one of the most important loci of lead-mediated toxicity, plays a key role in high cognitive functions, such as learning and memory (Squire, 1992). Long-term potent

14、iation (LTP) in the hippocampus is a form of activity-dependent synaptic plasticity that may be the electrophysiological substrate for learning and memory (Bliss and Lomo, 1973; Bliss and Collingridge, 1993). Pb-induced impairments of synaptic plasticity both in slices of hippocampus from Pb-exposed

15、 animals (Altmann et al., 1993) and in CA1 and dentate gyrus (DG) area of hippocampus in Pb-exposed rats in vivo (Gilbert et al., 1996, 1999; Zaiser and tic, 1997) have been reported. Our previous studies also showed that chronic Pb2 exposure impaired LTP in DG area of hippocampus in rats in vivo, w

16、hich possibly is associated with chronic lead-induced deficits of learning and memory in rats (Ruan et al., 1998).Although common chelating agents, such as triethyl-ene tetramine (TETA), penicillamine and desferrioxamine, are currently available for the treatment of Pb2 neurotox- icity, they are sho

17、wn to have many adverse effects and to be incapable of alleviating some neurotoxic effects of Pb2 . So far, no efficient drugs are available for treating chronic lead-induced deficits, especially, related to learn- ing and memory (Mortensen and Walson, 1993; Porru and Alessio, 1996). CQ, cliool (5-c

18、hloro-7-iodo-8-hydroxy- oline) was first prepared in the early part of the last century and was widely used as an biotic. In the 1970s, it was banned in many countries due to being linked to outbreak of subacute myelo-optic neuropathy (SMON) in Japan (Arbiser et al., 1998; Tateishi, 2000; Tabira, 20

19、01). However, a causal relation between CQ and SMON was never proven (Bush and Masters, 2001). In fact, CQ is a transition metal ion chelator and/or ionophore with low affi for divalent cations, such as: CQ for Zn2 (K1 7.0)0306-4522/07$30.00 0.00 2007 IBRO. Published by Elsevier . . doi:10.1016/j.ne

20、uroscience.2007.04.042853W.-H. Chen et al. / Neuroscience 147 (2007) 853 864863and CQ for Cu2 (K1 8.9) (Cherny et al., 2001) and it is hydrophobic and ly permeable across the blood brain barrier (Padmanabhan et al., 1989). Recently, as a novel and potential therapeutic strategy, CQ treatment has yie

21、lded some promising results in animal ms and clin- ical trials related to dysfunctions of metal ions. For exam- ple, it enhanced A ( -amyloid protein) aggregate disso- lution and decreased A toxicity in an Alzheimers disease (AD) m, where Cu and Zn are elevated in the neocor- tex and particularly co

22、ncentrated in amyloid plaques (Cherny et al., 2001); the phase II clinical trial showed that it displayed early positive effects on AD and had no signif- icant CQ neurotoxicity (Ritchie et al., 2003); it prevented MPTP (1-methyl-4-phenyl-1,2,3,6-tetra-pyridine)-induced neurotoxicity in vivo, a Parki

23、nsons disease (PD) m, where Fe levels in substa nigra (SN) have been re- ported to be elevated (Kaur et al., 2003); it decreased polyQ-expanded protein accumulation and promoted cell survival in an in vitro m of Huntingtons disease (HD) and decreased symptoms and increased lifespan in an animal m of

24、 HD, which is associated with dysfunctions of Cu, Fe or Zn (Nguyen et al., 2005). At the same time, CQ-associated neurological side effects, which may be due to a CQ chelating effect on the vitamin B12 (VB12)- bound Co2 , are now believed to be preventable with VB12 supplementation (Yassin et al., 2

25、000). VB12 plays a key role in CNS function. It participates in the methionine- synthase-mediated conversion of homocysteine to methi- onine, which is essential for nucleotide synthesis and genomic and non-genomic methylation (Reynolds, 2006). To determine whether CQ treatment could rescue the lead-

26、induced impairments of synaptic plasticity related to learning and memory, in the present study, we investigated the LTP of different treatment groups in hippocampal DG area of the anesthetized rats in vivo. In addition, we also investigated the input/output functions (I/Os) and paired- pulse reacti

27、ons (PPRs), two auxiliary electrophysiological parameters, and measured Pb2 concentrations in hip- pocampus of different treatment groups. Our results indi- cate: CQ alone does not rescue the lead-induced impair- ments of synaptic plasticity in hippocampal DG area of the anesthetized rats in vivo; V

28、B12 alone partly rescues the lead-induced impairments of LTP; however CQ and VB12 synergistically, totally, rescue these impairments in hip- pocampal DG area of the anesthetized rats in vivo and moreover, the effects of CQ and VB12 co-administrationare specific to the lead-exposed animals.EXPERIMENT

29、AL PROCEDURESExperimental animals and treatmentIn the present protocol, the rats were divided into five groups which included control group (control) (n 12), lead-exposed group (lead) (n 12), lead-exposed with cliool group (lead CQ) (n 10), lead-exposed with vitamin B12 group (lead VB12) (n 9), lead

30、-exposed with clio ol and vitamin B12 group (lead CQ VB12) (n 10) and control with cliool and vitamin B12 group (control CQ VB12) (n 8). Before pups delivery, all the mother rats ly had access to tap water. From pups delivery to wean- ing, the offspring had access either to tap water (the controls a

31、ndcontrol CQ VB12 group) or to water with 0.2% lead acetate (the lead-exposed group and groups related to the lead-exposed) via their mothers milk. After weaning, the offspring were weaned to the same solution as that given their mothers, so that chronic exposure to Pb2 was throughout the lifetime t

32、hat began in the early postnatal period. Before the experiment, the rats of drug- treatment group were intraperitoneally injected with the different agents for the distinct groups for 7 days. The doses of agents were 30 mg/kg/day for CQ and 45 g/kg/day for VB12 (Cherny et al., 2001). The experiments

33、 were carried out on adult Wistar rats weighing 200 280 g. At the age of 60 80 postnatal days, the animals were utilized for extracellular recording in aG area of hippocampus of anesthetized Wistar rats in vivo. No more than two animals per litter were utilized for experimental measurement in the sa

34、me group. The agents used in this experiment were all bought from Sigma (St. Louis, MO, USA). The care and use of animals in these experiments followed the guidelines and protocol approved by the Care and Use of Animals Committee of the University of Science and Technology of . All efforts were to m

35、inimize the number of animals used and their suffering. All experiments conformed to the U.S. National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH publica- tion No. 80-23, revised 1996).Stimulation and recordingIn each recording session, rats were anesthetized with ure

36、thane (1.8 g/kg, i.p. injection) and fixed in a stereotaxic head-holder. The skull was exposed and the animals body temperature, heart rate and electrocardiogram were monitored. A concentric bipolar stim- ulating electrode was placed in the lateral perforant path (coordi- nates with the skull surfac

37、e flat: 8.0 mm posterior to bregma, 4.2 mm lateral to the midline, and 2.8 3.0 mm below the surface of the skull). A micropipette glass recording electrode (35 m tip diameter and 13 M ) was placed in the DG (coordi- nates with the skull surface flat: 3.8 mm posterior to bregma, 2.1 mm lateral to the

38、 midline, and 3.0 3.5 mm below the surface of the skull). The glass micropipette filled with 2 M NaCl was used for extracellular recordings.I/OI/O curves were generated by systematic variation of the stimulus current by steps of 0.1 mA (0.11.0 mA) in order to evaluate synaptic potency. Stimulus puls

39、es were delivered at 0.05 Hz and three responses at each current level were averaged.PPRPPR was evaluated by increasing the interpulse intervals (IPIs, 10 400 ms). The stimulus current intensity was adjusted at in- tensity yielding 40 60% of the al amplitude of population spike (PS). Stimulus pairs

40、were delivered at 0.05 Hz and three responses were averaged at each IPI.LTPIn present study, LTP was recorded in each animal. After 20 min baseline recordings, a high frequency stimulus (HFS) was applied (250 Hz, 1 s). Posttetanic recordings were performed for 1 h with single pulse applied at a freq

41、uency of 0.05 Hz. At the end of each recording session, small electrolytic lesions (10 A, 10 s) were to permit histological verification of the tip position of the electrodes. Hippocampus was isolated for measuring Pb2 con- centrations.Hippocampal Pb2 concentrationsHippocampal Pb2 concentrations wer

42、e estimated on the animals used for electrophysiology. After decapitation of the animals, thehippocampi were isolated, and digested with an organic tissue solubilizer. The Pb2 concentrations of the hippocampus were measured by PlasmaQuad 3 inductive coupled plasma mass spectroscopy (VG Elemental , C

43、heshire, UK).Data analysisResponses were evoked by stimulating perforant path at low frequency (0.05 Hz). For each time-point, 20 evoked responses for LTP were averaged. Both field excitatory postsynaptic potential (fEPSP) slope and PS amplitude were monitored. fEPSP slope was measured as theal slop

44、e through the five steepest points obtained on the first positive deflection of the potential. The amplitude of PS was measured from the peak of the first positive deflection of the evoked potential to the peak of the following negative potential. All potentials employed as baseline criteria were ev

45、oked at a stimulus intensity which produces 40 60% of the al amplitude of PS. Mean values of LTP are presented as the mean S.E.M. Mean values were normalized to pre-tetanus baseline values. The amplitudes of LTP were calculated by aver- aging the percentage of post-tetanus data in 1 h compared with

46、pre-tetanus baseline data. Comparisons between each two given current levels or each two given IPIs or each two given time-points were analyzed by one-way ANOVA. Comparisons between each two groups were analyzed by two-way ANOVA with Tukey test. Probabilities less than 0.05 were considered as signif

47、icantly dif- ference.RESULTSCQ alone does not rescue the lead-induced impairments of synaptic plasticity in hippocampal DG area of the anesthetized rats in vivoConsistent with our previous report (Ruan et al., 1998), chronic Pb2 exposure induced impairments of synaptic plasticity in hippocampal DG a

48、rea of the anesthetized rats in vivo as shown in Fig. 1. The I/O curves were measured by fEPSP slope and PS amplitude. Compared with con- trols, the lead-exposed group had no significant difference in both the fEPSP slope (P 0.05) (Fig. 1A) and PS am- plitude (P 0.05) (Fig. 1B). This result suggests

49、 that chronic Pb2 exposure does not affect the basal synaptic transmission of dentate granule cells. In this experiment, the PPR of PS amplitude was measured by the second pulse stimulus-induced PS amplitude to the first pulse stimulus-induced PS amplitude ratio. The average peak facilitation was 22

50、5 17% in controls at IPI 60 ms and was significantly decreased to 180 13% in lead-exposed group at IPI 80 (P 0.05). Moreover, the corresponding average facilitations at IPI 40 ms (P 0.05), IPI 60 ms (P 0.05) and IPI 100 (P 0.05) ms were also significantly decreased in lead-exposed group compared wit

51、h controls (Fig. 1C and Fig. 5). These results demonstrate that chronic Pb2 exposure impairs the average peak facilita- tion of PPR and the corresponding average facilitations at some IPI points. As to LTP, in controls, the LTP amplitude was 136 4% (fEPSP slope) and 266 17% (PS ampli- tude), and was

52、 depressed significantly to 122 2% (fEPSP slope: F(1,22) 77.89, P 0.001) and 195 10% (PS ampli- tude: F(1,22) 118.32, P 0.001) in the lead-exposed group (Fig. 1E, F and Fig. 5). This result suggests that chronic Pb2 exposure impairs the induction of LTP of both fEPSP slope and PS amplitude in DG are

53、a of hippocampus.To determine whether CQ could repair these impair- ments in lead-exposure groups, we investigated the effects of CQ on chronic Pb2 exposure by 1-week CQ injectioni.p. First, compared with the lead-exposed group, the lead CQ group had no significant difference in the PS amplitude of

54、I/O curves (P 0.05) (Fig. 1B), but had a significant decrease in the fEPSP slope of I/O curves (F(1,20) 10.30, P 0.05) (Fig. 1A). At the same time, the fEPSP slope of I/O curves from the lead CQ group were significantly decreased as compared with controls (fEPSP slope: F(1,20) 10.13, P 0.05) (Fig. 1

55、A). These results in- dicate that the lead CQ group inhibits the fEPSP slope of basal synaptic transmission. Second, the PPR of PS am- plitude had no significant difference in the average peak facilitation and the corresponding average facilitations/ depressions at all IPI points between the lead CQ

56、 group and lead-exposed group (P 0.05) (Fig. 1D and Fig. 5). Third, the LTP amplitudes of both fEPSP slope and PS amplitude also had no significant difference between the lead CQ group and lead-exposed group (P 0.05 for both fEPSP slope and PS amplitude) (Fig. 1E, F and Fig. 5).Taken together, chron

57、ic Pb2 exposure induces im- pairments of PPR and LTP in hippocampal DG area of the anesthetized rats in vivo. However, CQ alone does not rescue these impairments. Furthermore, it seems to inhibit the fEPSP slope of basal synaptic transmission.VB12 partly rescues the lead-induced impairments of synap

58、tic plasticity in the hippocampal DG area of the anesthetized rats in vivoNext, we investigated the effects of VB12 on the lead- induced impairments in the hippocampal DG area of the anesthetized rats in vivo. Though the PS amplitude of I/O curves from the lead VB12 group was significantly de- creas

59、ed as compared with that from controls, but com- pared with the lead-exposed group, the lead VB12 group had no significant difference in both the fEPSP slope (P 0.05) and PS amplitude of I/O curves (P 0.05) (Fig. 2A and B). The PPR of PS amplitude had no significant difference in the average peak facilitation and the corre- sponding average facilitations/depressions at all IPI