Molecular Brain - Latest Articles

Rapid synaptic potentiation within the anterior cingulate cortex mediates trace fear learning

Giannina Descalzi — 2012-02-03T00:00:00Z

Although the cortex has been extensively studied in long-term memory storage, less emphasis has been placed on immediate cortical contributions to fear memory formation. AMPA receptor plasticity is strongly implicated in learning and memory, and studies have identified calcium permeable AMPA receptors (CP-AMPARs) as mediators of synaptic strengthening. Trace fear learning engages the anterior cingulate cortex (ACC), but whether plastic events occur within the ACC in response to trace fear learning, and whether GluN2B subunits are required remains unknown. Here we show that the ACC is necessary for trace fear learning, and shows a rapid 20% upregulation of membrane AMPA receptor GluA1 subunits that is evident immediately after conditioning. Inhibition of NMDA receptor GluN2B subunits during training prevented the upregulation, and disrupted trace fear memory retrieval 48 h later. Furthermore, intra-ACC injections of the CP-AMPAR channel antagonist, 1-naphthylacetyl spermine (NASPM) immediately following trace fear conditioning blocked 24h fear memory retrieval. Accordingly, whole cell patch clamp recordings from c-fos positive and c-fos negative neurons within the ACC in response to trace fear learning revealed an increased sensitivity to NASPM in recently activated neurons that was reversed by reconsolidation update extinction. Our results suggest that trace fear learning is mediated through rapid GluN2B dependent trafficking of CP-AMPARs, and present in vivo evidence that CP-AMPAR activity within the ACC immediately after conditioning is necessary for subsequent memory consolidation processes.

Hippocampal function is not required for the precision of remote place memory

Takashi Kitamura — 2012-02-02T00:00:00Z

Background: During permanent memory formation, recall of acquired place memories initially depends on the hippocampus and eventually become hippocampus-independent with time. It has been suggested that the quality of original place memories also transforms from a precise form to a less precise form with similar time course. The question arises of whether the quality of original place memories is determined by brain regions on which the memory depends. Results: To directly test this idea, we introduced a new procedure: a non-associative place recognition memory test in mice. Combined with genetic and pharmacological approaches, our analyses revealed that place memory is precisely maintained for 28 days, although the recall of place memory shifts from hippocampus-dependent to hippocampus-independent with time. Moreover, the inactivation of the hippocampal function does not inhibit the precision of remote place memory. Conclusion: These results indicate that the quality of place memories is not determined by brain regions on which the memory depends.

In vivo antinociception of potent mu opioid agonist tetrapeptide analogues and comparison with a compact opioid agonist - neurokinin 1 receptor antagonist chimera

Karel Guillemyn — 2012-01-30T00:00:00Z

Background: An important limiting factor in the development of centrally acting pharmaceuticals is the blood-brain barrier (BBB). Transport of therapeutic peptides through this highly protective physiological barrier remains a challenge for peptide drug delivery into the central nervous system (CNS). Because the most common strategy to treat moderate to severe pain consists of the activation of opioid receptors in the brain, the development of active opioid peptide analogues as potential analgesics requires compounds with a high resistance to enzymatic degradation and an ability to cross the BBB. Results: Herein we report that tetrapeptide analogues of the type H-Dmt-Xxx-Yyy-Gly-NH2 are transported into the brain after intravenous and subcutaneous administration and are able to activate the mu and delta opioid receptors more efficiently and over longer periods of time than morphine. Using the hot water tail flick test as the animal model for antinociception, a comparison in potency is presented between a side chain conformationally constrained analogue containing the benzazepine ring (BVD03, Yyy: Aba), and a "ring opened" analogue (BVD02, Yyy: Phe). The results show that in addition to the increased lipophilicity through amide bond N-methylation, the conformational constraint introduced at the level of the Phe side chain causes a prolonged antinociception. Further replacement of NMe-D-Ala by D-Arg in the tetrapeptide sequence led to an improved potency as demonstrated by a higher and maintained antinociception for AN81 (Xxx: D-Arg) vs. BVD03 (Xxx: NMe-D-Ala). A daily injection of the studied opioid ligands over a time period of 5 days did however result in a substantial decrease in antinociception on the fifth day of the experiment. The compact opioid agonist - NK1 antagonist hybrid SBCHM01 could not circumvent opioid induced tolerance. Conclusions: We demonstrated that the introduction of a conformational constraint has an important impact on opioid receptor activation and subsequent antinociception in vivo. Further amino acid substitution allowed to identify AN81 as an opioid ligand able to access the CNS and induce antinociception at very low doses (0.1 mg/kg) over a time period up to 7 hours. However, tolerance became apparent after repetitive i.v. administration of the investigated tetrapeptides. This side effect was also observed with the dual opioid agonist-NK1 receptor antagonist SBCHM01.

Genetic marking and characterization of Tac2-expressing neurons in the central and peripheral nervous system

Lynn Mar — 2012-01-24T00:00:00Z

Background: The neurocircuits that process somatic sensory information in the dorsal horn of the spinal cord are still poorly understood, with one reason being the lack of Cre lines for genetically marking or manipulating selective subpopulations of dorsal horn neurons. Here we describe Tac2-Cre mice that were generated to express the Cre recombinase gene from the Tac2 locus. Tachykinin 2 (Tac2) encodes a neurotransmitter, neurokinin B (NKB). Results: By crossing Tac2-Cre mice with ROSA26-tdTomato reporter mice, we directly visualized Tac2 lineage neurons in the dorsal root ganglia, the dorsal horn of the spinal cord, and many parts of the brain including the olfactory bulb, cerebral cortex, amygdala, hippocampus, habenula, hypothalamus, and cerebellum. This Tac2-Cre allele itself was a null allele for the Tac2 gene. Behavioral analyses showed that Tac2 homozygous null mice responded normally to a series of algogenic (pain-inducing) and pruritic (itch-inducing) stimuli. Conclusions: Tac2-Cre mice are a useful tool to mark specific subsets of neurons in the sensory ganglia, the dorsal spinal cord, and the brain. These mice can also be used for future genetic manipulations to study the functions of Tac2-expressing neurons or the functions of genes expressed in these neurons.

Understanding the physiological roles of the neuronal calcium sensor proteins

Robert Burgoyne — 2012-01-23T00:00:00Z

Calcium signalling plays a crucial role in the control of neuronal function and plasticity. Changes in neuronal Ca2+ concentration are detected by Ca2+-binding proteins that can interact with and regulate target proteins to modify their function. Members of the neuronal calcium sensor (NCS) protein family have multiple non-redundant roles in the nervous system. Here we review recent advances in the understanding of the physiological roles of the NCS proteins and the molecular basis for their specificity.

NMDA Receptor Activation Stimulates Transcription-Independent Rapid Wnt5a Protein Synthesis via the MAPK Signaling Pathway

Yichen Li — 2012-01-04T00:00:00Z

Wnt proteins are emerging key regulators of the plasticity and functions of adult brains. However, the mechanisms by which the expression of Wnt proteins is regulated in neurons are unclear. Using primary cortical cultures, we show that activation of NMDA receptors (NMDARs) induces rapid Wnt5a protein synthesis. This NMDAR-regulated Wnt5a synthesis does not require transcription and is a result of activity-dependent translation. We also show that NMDAR-regulated Wnt5a translation depends on MAPK signaling but not mTOR signaling. Our findings suggest that the synaptic activity of CNS neurons activates NMDARs, which in turn stimulate translation from stored Wnt5a mRNA via the MAPK signaling pathway.

Dependence of NMDA/GSK3beta Mediated Metaplasticity on TRPM2 Channels at Hippocampal CA3-CA1 Synapses

Yu-Feng Xie — 2011-12-21T00:00:00Z

Transient receptor potential melastatin 2 (TRPM2) is a calcium permeable non-selective cation channel that functions as a sensor of cellular redox status. Highly expressed within the CNS, we have previously demonstrated the functional expression of these channels in CA1 pyramidal neurons of the hippocampus. Although implicated in oxidative stress-induced neuronal cell death, and potentially in neurodegenerative disease, the physiology of transient receptor potential melastatin 2 channels (TRPM2) is unknown. Interestingly, we have shown that the activation of these channels may be sensitized by co-incident NMDA receptor activation, suggesting a potential contribution of TRPM2 to synaptic transmission. Using hippocampal cultures and slices from TRPM2 null mice we demonstrate that the loss of these channels selectively impairs NMDAR-dependent long-term depression (LTD) while sparing long-term potentiation. Impaired LTD resulted from an inhibition of GSK3beta, through increased phosphorylation, and a reduction in the expression of PSD95 and AMPARs. Notably, LTD could be rescued in TRPM2 null mice by recruitment of GSK3beta signalling following dopamine D2 receptor stimulation. We propose that TRPM2 channels play a key role in hippocampal synaptic plasticity.

Neuromelanin, neurotransmitter status and brainstem location determine the differential vulnerability of catecholaminergic neurons to mitochondrial DNA deletions

Matthias Elstner — 2011-12-21T00:00:00Z

Background: Deletions of the mitochondrial DNA (mtDNA) accumulate to high levels in dopaminergic neurons of the substantia nigra pars compacta (SNc) in normal aging and in patients with Parkinson's disease (PD). Human nigral neurons characteristically contain the pigment neuromelanin (NM), which is believed to alter the cellular redox-status. The impact of neuronal pigmentation, neurotransmitter status and brainstem location on the susceptibility to mtDNA damage remains unclear. We quantified mtDNA deletions in single pigmented and non-pigmented catecholaminergic, as well as non-catecholaminergic neurons of the human SNc, the ventral tegmental area (VTA) and the locus coeruleus (LC), using laser capture microdissection and single-cell real-time PCR. Results: In healthy aged individuals, mtDNA deletion levels were highest in pigmented catecholaminergic neurons (25.2 +/- 14.9%), followed by non-pigmented catecholamergic (18.0 +/- 11.2%) and non-catecholaminergic neurons (12.3 +/- 12.3%; p < 0.001). Within the catecholaminergic population, mtDNA deletion levels were highest in dopaminergic neurons of the SNc (33.9 +/- 21.6%) followed by dopaminergic neurons of the VTA (21.9 +/- 12.3%) and noradrenergic neurons of the LC (11.1 +/- 11.4%; p < 0.001). In PD patients, there was a trend to an elevated mutation load in surviving non-pigmented nigral neurons (27.13 +/- 16.73) compared to age-matched controls (19.15 +/- 11.06; p = 0.052), but levels where similar in pigmented nigral neurons of PD patients (41.62 +/- 19.61) and controls (41.80 +/- 22.62). Conclusions: Catecholaminergic brainstem neurons are differentially susceptible to mtDNA damage. Pigmented dopaminergic neurons of the SNc show the highest levels of mtDNA deletions, possibly explaining the exceptional vulnerability of the nigro-striatal system in PD and aging. Although loss of pigmented noradrenergic LC neurons also is an early feature of PD pathology, mtDNA deletion levels are not elevated in this nucleus in healthy controls. Thus, mtDNA deletions are neither an inevitable consequence of catecholamine metabolism nor an universal explanation for the regional vulnerability seen in PD.

The amount of astrocytic GABA positively correlates with the degree of tonic inhibition in hippocampal CA1 and cerebellum

Bo-Eun Yoon — 2011-11-22T00:00:00Z

A tonic form of synaptic inhibition occurs in discrete regions of the central nervous system and has an important role in controlling neuronal excitability. Recently, we reported that GABA present in astrocyte is the major source of tonic inhibition in cerebellum and that GABA is released through Bestrophin-1 channel by direct permeation. In this study, we screened for the presence of astrocytic GABA in various brain regions such as hippocampus, thalamus, hypothalamus and cerebellum using immunohistochemistry. We found that astrocytic GABA was present in the regions that were reported to show tonic inhibition. Because the existence of tonic inhibition in hippocampal CA1 is somewhat controversial, we compared the amount of astrocytic GABA and tonic inhibition between the hippocampal CA1 pyramidal cell layer and the cerebellar granule cell layer. Unlike cerebellar glial cells, hippocampal astrocytes did not contain GABA. The tonic inhibition was also much lower in the pyramidal neurons of hippocampal CA1 compared to the granule cells of cerebellum. Nevertheless, most of the hippocampal astrocytes expressed Bestrophin-1 channel. These data indicate that the absence of astrocytic GABA results in a low level of tonic inhibition in hippocampal CA1 region.

Slice orientation and muscarinic acetylcholine receptor activation determine the involvement of N-methyl D-aspartate receptor subunit GluN2B in hippocampal area CA1 long-term depression

Thomas Bartlett — 2011-11-15T00:00:00Z

Background: The contribution of different GluN2 subunits of the N-methyl D-aspartate (NMDA) receptor to the induction of bidirectional hippocampal synaptic plasticity is a controversial topic. As both supporting and refuting evidence for the hypothesis of subunit specialization in opposing directions of plasticity has accumulated since it was first proposed a few years ago, we hypothesize that differences in experimental conditions may have in part contributed to some of the inconsistent results from these studies. Here we investigate the controversial hypothesis that long-term depression (LTD) is preferentially induced by GluN2B-containing NMDA receptors in area CA1 of hippocampal slices. Results: We find that brain slices from 2-3 week old rats prepared in the sagittal orientation have GluN2B-independent LTD whereas slices prepared in the coronal orientation have GluN2B-dependent LTD. There was no difference between the orientations in the fraction of the NMDAR EPSC sensitive to a GluN2B-selective antagonist, leading us to believe that the intracellular signaling properties of the NMDARs were different in the two preparations. Coronal slices had greater association of LTD-related intracellular signaling protein RasGRF1 with GluN2B relative to sagittal slices. Antagonism of muscarinic acetylcholine receptors (mAChRs) in the sagittal slices returned LTD to a GluN2B-dependent form and increased the association of GluN2B with RasGRF1. Conclusions: These results suggest a novel form of NMDAR modulation by mAChRs and clarify some disagreement in the literature.