Selective Cholinergic Depletion in Medial Septum Leads to Impaired Long Term Potentiation and Glutamatergic Synaptic Currents in the Hippocampus

Cholinergic depletion in the medial septum (MS) is associated with impaired hippocampal-dependent learning and memory. Here we investigated whether long term potentiation (LTP) and synaptic currents, mediated by alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA) and N-methyl-D-aspartate (NMDA) receptors in the CA1 hippocampal region, are affected following cholinergic lesions of the MS. Stereotaxic intra-medioseptal infusions of a selective immunotoxin, 192-saporin, against cholinergic neurons or sterile saline were made in adult rats. Four days after infusions, hippocampal slices were made and LTP, whole cell, and single channel (AMPA or NMDA receptor) currents were recorded. Results demonstrated impairment in the induction and expression of LTP in lesioned rats. Lesioned rats also showed decreases in synaptic currents from CA1 pyramidal cells and synaptosomal single channels of AMPA and NMDA receptors. Our results suggest that MS cholinergic afferents modulate LTP and glutamatergic currents in the CA1 region of the hippocampus, providing a potential synaptic mechanism for the learning and memory deficits observed in the rodent model of selective MS cholinergic lesioning

Modulatory effects of dextran sulfate and fucoidan on binding and channel properties of AMPA receptors isolated from rat brain

Previous work showed that the glycosaminoglycan (GAG) dextran sulfate (500 kDa) altered the binding and channel properties of α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA)‐type glutamate receptors. The current study compared the effects of dextran sulfate with another GAG, fucoidan (100–180 kDa), to determine whether GAG‐mediated changes in high‐affinity binding of AMPA receptors have a concomitant influence on specific channel properties. Dextran sulfate was more potent in inhibiting high‐affinity AMPA binding to solubilized receptors (EC50 of 7 nM) compared to fucoidan (EC50 of 124 nM). Similarly, dextran sulfate was more potent in modulating the channel properties of purified and reconstituted AMPA receptors. Dextran sulfate, at 1 μg/ml (2 nM), produced a three to fourfold increase in open channel probability and a threefold increase in mean burst duration of channel activity elicited by 283 nM AMPA. The mean open time was increased by two to threefold and closed times were decreased by two to eightfold. Fucoidan produced similar effects at a concentration many times higher than that of dextran sulfate. Dextran sulfate and fucoidan had no effect on the single channel conductance or the ability of a specific antagonist to block AMPA channels. The effects of GAGs on multichannel patches showed an interactive channel gating behavior resulting in macroscopic currents with long lived open channel life times. These findings suggest that GAG components of proteoglycans can interact with and alter the binding affinity of AMPA receptors and modulate their functional properties. Synapse 60:456–464, 2006. © 2006 Wiley‐Liss, Inc

D-Galactose Effectiveness in Modeling Aging and Therapeutic Antioxidant Treatment in Mice

Accumulating evidence suggests that mitochondrial dysfunction and oxidative stress play major roles in aging. Chronic administration of D-galactose has been reported to cause deterioration of cognitive and motor skills that are similar to symptoms of aging and, therefore, is regarded as a model of accelerated aging. Because enhancing endogenous antioxidants is now widely regarded as an attractive therapy for conditions associated with mitochondrial oxidative stress, in the present study the effects of α-lipoic acid, L-carnitine, and PMX-500F on D-galactose treated mice were tested. Female mice were injected with (100 mg/kg) D-(+)-galactose for 6 weeks and some groups were treated with a daily dose of α-lipoic acid (5 mg/kg), L-carnitine (3.9 mg/kg), PMX-500F (11.9 mg/kg), or the vehicle (0.1 M Tris, pH 7.4). Control mice were treated with physiological saline. An accelerating Rota-Rod, open field test, and Y-maze test were performed, and serum lactate concentrations were analyzed. These analyses did not identify impairment in motor coordination, open-field activity, or spatial memory (p > 0.05). Similarly, serum lactate concentrations in D-galactose-treated mice were not elevated when compared to controls (p > 0.05). Treatment with the antioxidant compounds at the given concentrations did not result in any changes in the behavioral parameters tested. In conclusion, results of this study illustrate that chronic, short-term D-galactose treatment may not represent a suitable model for inducing readily detectable age-related neurobehavioral symptoms in mice

Impairment of LTP in 192-saporin infused rats.

<p>(A) Summary data for experiments in which LTP was induced by TBS and measured at 55–60 min after TBS. LTP in 192-saporin infuced rats (gray circles) was reduced compared to the saline infused controls (black circles). (B) Sample traces depicting LTP in saline infused control rats and lack of LTP in 192-saporin infused rats. Traces with gray lines represent those collected prior to TBS, during baseline recording, and the traces with black lines represent those taken 55–60 min after TBS. Calibration: 1 mV, 20 ms. (c) Bar chart showing drastic reduction of LTP in 192-saporin infused rats (gray) compared to the saline infused controls (black).</p

Cholinergic depletion in the MS leads to alterations in NMDA elicited synaptic single channel currents.

<p>Sample traces and respective current amplitude histograms of (A) non-infused control rats, (B) saline infused rats, and (C) 192-saporin infused rats show that channel open peak (right peaks in each histogram) in the 192-saporin data is reduced demonstrating a decrease in open probability (Calibration: each 4 pA, 200 ms). Dwell open time histograms of NMDA currents were best fitted with two terms for (D) non-infused, (E) saline infused, and (F) 192 saporin infused data. The NMDA elicited currents were confirmed by addition of APV to the extracellular solution to completely block these currents (Data not shown).</p

Inhibition of MS cholinergic pathway decreases the AMPA receptor activity in the CA1 pyramidal neurons of the hippocampus.

<p>(A) Representatives of AMPA receptor mediated sEPSC traces in controls and 192-saporin infused rats recorded at −65 mV membrane potential; Calibration: 40 pA, 1 s. The adjacent average traces depict the reduction in the amplitude in the MS cholinergic lesioned rats compared to control rats, which show no difference in amplitude; Calibration: 15 pA, 30 ms. (B) Cumulative fraction plot of sEPSC amplitude from the composite data shows the shift of 192-saporin curve to the left from the controls, indicating reductions in amplitude. (C) Cumulative fraction plots of sEPSC interevent intervals exhibiting increased values for 192-saporin infused rats suggesting decreased frequency. (D) Representative traces of AMPA receptor mediated mEPSCs for the three groups (Calibration: 10 pA, 1 s) and the mEPSCs (Calibration: 10 pA, 30 ms) to show reduced amplitude in the 192-saporin infused rats. (E) Cumulative fraction plots for amplitude and (F) interevent interval exhibiting reduced amplitude and frequency.</p

Burst analysis of AMPA and NMDA receptor single channel currents in synaptosomes.

<p>*<i>P</i><0.05,</p>#<p><i>P</i><0.01.</p

Lesioning of MS cholinergic pathway decreases the NMDA receptor mediated currents in the CA1 pyramidal neurons.

<p>(A) Representatives of NMDA receptor mediated sEPSC traces recorded at −40 mV membrane potential shows reduction of the frequency in the 192-saporin infused rats compared to the controls (Calibration: 40 pA, 1 s). The adjoining average traces of sEPSCs exhibit the reduction in amplitude in the 192-saporin infused rats (Calibration: 15 pA, 30 ms). Cumulative fraction curves of (B) amplitude and (C) interevent interval showing reduction in amplitude and frequency in the 192-saporin infused rats. (D) Representative segments of NMDA mediated mEPSCs (Calibration: 10 pA, 1 s) and average mEPSCs (Calibration: 10 pA, 30 ms) depicts reduced amplitude and frequency in the 192-saporin infused rats compared to the two controls. Cumulative fraction curves for (E) amplitude and (F) interevent interval of mEPSCs.</p

Infusion of 192-saporin results in lesioning of cholinergic neurons.

<p>Photomicrographs (50× magnification) of MS region immunostained with anti-ChAT antibody in (A) a saline infused rat and (B) in a rat infused with 192-saporin.</p