Increased susceptibility to LTP generation and changes in NMDA-NR1 and -NR2B subunits mRNA expression in rat hippocampus after MCH administration

doi:10.1016/j.peptides.2003.09.006

Peptides

Volume 24, Issue 9, September 2003, Pages 1403-1411

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Abstract

The present study attempts to determine which mechanisms underlie the retrograde facilitation of memory induced by microinjection hippocampal melanin-concentrating hormone (MCH) on the inhibitory avoidance paradigm. Previous reports using this test on the hippocampus suggest that NMDA receptor-mediated mechanisms are involved in memory processing and are also necessary for the induction of long-term potentiation (LTP) of the hippocampal dentate gyrus. In addition, alterations in expression of synaptic NMDA subunits in the hippocampus have been associated with memory formation of an inhibitory avoidance task. We have studied the effects of the neuropeptide upon the electrophysiological parameters using hippocampal slices from rats injected with the peptide and tested in step-down tests as well as possible changes in the mRNA expression of NMDA receptor subunits. We postulate that the increased facility to induce LTP, and the overexpression of this N-methyl-d-aspartate mRNA receptor subunits induced by MCH, could be behind the retrograde facilitation observed after MCH hippocampal microinjection.

Introduction

Many studies have demonstrated that the hippocampus is involved in the formation and in the expression of memory for one-trial step-down inhibitory avoidance task in rats [3], [4], [36], [56] and is an essential structure for the memory and learning processes [2], [40], [45], [58]. This is the task most often used for research on the mechanisms of memory formation and modulation because it is learned very rapidly and memory can be easily and reliably assessed for days, even or months following training. The use of this learned task in hippocampus has helped to identify the major modulatory and core mechanisms of memory formation, and the biochemical sequence of events for memory formation are now well known [17], [18], [19]. The sequence initially involves the activation of different types of glutamate receptors: the a-amino-3-hydroxy-5-methyl-4-isoxazole propoinic acid (AMPA), and the N-methyl-d-aspartate (NMDA) receptors [9], [17], [20], [57]. The NMDA receptors are associated with the remarkable plasticity of the hippocampus [11], and are crucially involved in the neural mechanisms underlying certain forms of learning. In this sense, these receptors have also been shown to be necessary for the induction of long-term potentiation (LTP) in hippocampal dentate gyrus [8], [15]. This activation of glutamate receptors is followed by changes in second messengers and biochemical cascades led by enhanced activity of protein kinases A, C, and G and also calcium-calmodulin protein kinase II, and changes in glutamate receptor subunits and binding properties [18], [19]. The molecular steps that are related to memory formation of inhibitory avoidance behavior in the rat hippocampus agrees with the events described to LTP [18]. LTP is considered be a neural form of memory [28].

The NMDA receptor is composed of combinations of NR1, NR2A–D, and NR3 subunits, [13], [29], with NR1 subunit being a necessary component of functional NMDA receptor channels [29]; the loss of NR1 from CA1 cells results in a selective loss of NMDAR function without affecting other glutamate receptors (e.g. AMPA receptors). In addition, neither NMDA-dependent LTP nor LTD can be induced in the CA1 region and also results in spatial learning deficits. In addition, it has recently been demonstrated that alterations in expression of synaptic NMDA NR1 subunits in the hippocampus are associated with memory formation of an inhibitory avoidance task [5]. Coexpression of the NR1subunits with NR2 subunits enhances the individual functional properties of the subunits, such as channel conductance, along with agonist, antagonist, and co-agonist sensitivities [30], [35]. Tang et al. [55] have demonstrated that the overexpression of the recombinant NR1/NR2B subunits results in a prolonged opening of the NMDA receptor channel. In addition, an enhanced tyrosine phosphorylation of NR2B subunit has been observed following LTP induction “in vivo” [43], [44].

Melanin-concentrating hormone (MCH) is a cyclic peptide first isolated from the salmon pituitary [22]. It is subsequently found to be present in mammals [53] and MCH-producing neurons located mainly in the lateral hypothalamus and zona incerta, and projects into a wide variety of brain regions [1]. This extensive distribution suggests that MCH plays many physiological functions in the central nervous system. Recently, MCH research has actively focused on its roles in the regulation of feeding behavior and energy balance [21], [41], [47]. Besides this effect, MCH is also involved in other neuronal processes such as anxiety [31], [33], learning [32], [51], and reproduction [14], [50].

In our laboratory we have studied the effect of micro-infusion of MCH in hippocampus in the one-trial step-down inhibitory avoidance test because MCH contains cells that project into the hippocampus [1], and also, a high density of specific binding sites for the peptide have been found in this area [12].

We have shown that the micro-infusion of MCH in hippocampus increases the latency time in the one-trial step-down inhibitory test when injected immediately after training, facilitating the formation and retention of memory [32]. These changes are correlated with a significant increase in NO and cGMP hippocampal levels suggesting that MCH activates certain steps of the early biochemical memory cascade [51]. Furthermore, we have demonstrated in an electrophysiological model, that MCH addition to the perfusion chamber causes a long-lasting increase in the efficacy of the hippocampal synaptic transmission and that previous perfusion with MK-801 (a non competitive NMDA receptor antagonist) was able to impair the potentiation elicited by MCH [52].

Thus, in order to investigate the neurobiological events underlying the behavioral effects induced by MCH, in the present research we have studied the effects of the neuropeptide upon the electrophysiological parameters using hippocampal slices from rats injected with the peptide and tested in step-down tests, as well as possible changes in the mRNA expression of NMDA receptor subunits.

Section snippets

Animals

The experiments were performed on male Wistar rats of 230–280 g (age, 9–12 weeks) with all experiments having food and water ad libitum. The animals were housed in plastic cages (four or five in a cage) under a 12 h-light/12 h-dark cycle (lights on at 7.00 a.m.) at a constant temperature of 23±1 °C. The experimental sessions were conducted during the light phase of the cycle. Each animal was used in only one experiment.

Surgery

The animals were anaesthetized with chloral hydrate (400 mg/kg; i.p.) and placed

Behavioral and electrophysiological studies

The effects of intrahippocampal administration of acsf (control animals) or rMCH upon latency time are shown in Fig. 1. The infusion of MCH (0.5 μg/side) into the hippocampus immediately after training (0 h) caused a significant increase in the latency time with respect to the control animals (P<0.05 for Mann–Whitney U test).

Fig. 2 shows the threshold necessary to induce LTP in our electrophysiological model, measured in hertz. It can be seen that rats treated with rMCH showed an increased

Discussion

Our previous behavioral data showed that intrahippocampal MCH injection increases the latency time in the step-down test when is administered immediately after the training session, indicating that the peptide modifies memory retention [32].

In the present study we have confirmed these previous results (Fig. 1), and examined changes in hippocampal LTP after testing. The fact that MCH diminished the threshold in LTP generation indicates a positive correlation between this phenomenon and the MCH

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Increased susceptibility to LTP generation and changes in NMDA-NR1 and -NR2B subunits mRNA expression in rat hippocampus after MCH administration

Abstract

Introduction

Section snippets

Animals

Surgery

Behavioral and electrophysiological studies

Discussion

Neuron

Trends Neurosci.

Trends Pharmacol. Sci.

Trends Neurosci.

Mol. Brain Res.

Brain Res.

Brain Res.

Neurobiol. Learn. Mem.

Learn. Mem.

Behav. Neural. Biol.

Neuron

Physiol. Behav.

Peptides

Peptides

Biochem. Biophys. Res. Commun.

Behav. Brain Res.

Neurosci. Lett.

Behav. Brain Res.

Peptides

Peptides

The melanin-concentrating hormone system of rat brain: an immuno-and hybridization histochemical characterization

J. Comp. Neurol.

A synaptic model of memory: long-term potentiation in the hippocampus

Nature

Rapid and transient learning-associated increase in NMDA NR1 subunit in the rat hippocampus

Neurochem. Res.

Melanin-concentrating hormone is the cognate ligand for the orphan G-protein-coupled receptor SLC-1

Nature

Subtype-dependence of NMDA receptor channel open probability

J. Neurosci.

Excitatory amino acids in synaptic transmission in the Schaffer collateral-commissural pathway of the rat hippocampus

J. Physiol.

Anatomical organization of excitatory amino acids receptors and their pathways

Trends Neurosci.

Binding the sites for melanin-concentrating hormone (MCH) in brain synaptosomes and membranes from peripheral tissues identified with highly tritiated MCH

J. Recept. Signal Transduct. Res.

Stimulatory effect of melanin-concentrating hormone on luteinizing hormone release

Neuroendocrinology