Masao Maekawa

Spatial learning and long-term potentiation of mutant mice lacking d-amino-acid oxidase

We evaluated the role of D-amino-acid oxidase on spatial learning and long-term potentiation (LTP) in the hippocampus, since this enzyme metabolizes D-amino-acids, some of which enhance the N-methyl-D-aspartate receptor functions. The Morris water maze learning and the LTP in the CA1 area of the hippocampal slice were observed in wild-type mice and mutant mice lacking D-amino-acid oxidase. The mutant mice showed significantly shorter platform search times in the water maze and significantly larger hippocampal LTPs than the wild-type mice. These results suggest that the abundant D-amino-acids in the mutant mouse brain facilitate hippocampal LTP and spatial learning.

Facilitatory effects of subanesthetic sevoflurane on excitatory synaptic transmission and synaptic plasticity in the mouse hippocampal CA1 area

Behavioral investigations have shown that general anesthetics at low concentration have enhancing effects on learning and memory in some animal models. In the present experiments, in order to elucidate the cellular mechanisms underlying such memory enhancement, the effects of anesthetics at low doses on synaptic plasticity in the hippocampus were investigated. Tight-seal whole-cell recordings were made from CA1 pyramidal cells in hippocampal slices prepared from adult male mice, and the effects of subanesthetic concentrations of the volatile anesthetic sevoflurane on the glutamatergic excitatory postsynaptic currents (EPSCs) were investigated. In addition, extracellular recordings of field excitatory postsynaptic potential (fEPSP) and population spike (PS) were made, and the effects of subanesthetic sevoflurane on long-term potentiation (LTP) of the fEPSP slope and on LTP of PS amplitude were analyzed. Sevoflurane at anesthetic concentration inhibited the amplitude of EPSCs with an increase in the paired-pulse facilitation (PPF) ratio. In contrast, subanesthetic sevoflurane increased the amplitude of EPSCs without any appreciable changes in the PPF ratio. Subanesthetic sevoflurane also showed facilitatory influences on LTP of PS amplitude but not on LTP of the fEPSP slope. These observations suggest that sevoflurane at anesthetic concentration presynaptically inhibits excitatory synaptic transmission and at subanesthetic concentration postsynaptically enhances excitatory synaptic transmission in the hippocampal CA1 region. Further, subanesthetic sevoflurane seems to exert facilitatory effects on the EPSP-to-spike coupling process in the postsynaptic neurons. These results might provide clues as to the cellular mechanism of light level of sevoflurane anesthesia.

Difference in selectivity to song note properties between the vocal nuclei of the zebra finch

Responses to a single note from a zebra finch song and an artificially modified note were recorded from neurons in the vocal nuclei in the zebra finch. Response patterns recorded in the higher vocal center (HVC) and the lateral magnocellular nucleus of the neostriatum (IMAN) were compared. A subset of neurons were found to preferentially respond to the original temporal sequence of the components of the note and/or the specific harmonic structure of the note. These neurons were more common in the IMAN than in the HVC. These results suggest that response selectivity to both the harmonic and temporal pattern of the note increases as auditory information flows from the HVC to the IMAN.

Auditory responses in the nucleus basalis of the pigeon

Single-unit responses and field potentials evoked by sound stimuli were recorded from the nucleus basalis in the frontal lobe of the pigeon brain. Auditory units in this nucleus showed phasic responses to tonal stimuli and sharp frequency tuning. The latency of responses was 5.8 +/- 0.6 ms. The response properties of these units were similar to those of units in the nucleus mesencephalicus lateralis, pars dorsalis (a mesencephalic auditory nucleus), where fibers from the auditory nuclei in the medulla terminate. This suggests that the nucleus basalis receives projections from medullary auditory neurons. Averaged auditory field potentials in the frontal lobe had three prominent components. Regional changes in the polarity of each component suggest that auditory neurons in the medulla project directly to the nucleus basalis, which in turn projects to the frontal neostriatum.

Strategies for harmonic structure discrimination by zebra finches.

We trained zebra finches on a go/no-go operant conditioning procedure to discriminate between two harmonic structures that differed only by the presence or absence of the second harmonic. Test sessions revealed that the birds responded to probe stimuli based on the presence or absence of the second harmonic regardless of the other frequency components. Some of the birds also tended to use the fundamental frequency as a discriminative cue when the number of harmonics was few. These results indicate that birds use multiple strategies for this type of harmonic structure discrimination.

Differential expression of NMDA receptors in serotonergic and/or GABAergic neurons in the midbrain periaqueductal gray of the mouse

N-methyl-d-aspartate (NMDA) receptors expressed in the midbrain periaqueductal gray (PAG) exert various physiological functions. The PAG contains various neurotransmitter phenotypes, which include GABAergic neurons and serotonergic neurons. In the present experiments, we made tight-seal whole-cell recordings from GABAergic and/or serotonergic neurons in mouse PAG slices and analyzed NMDA and non-NMDA receptor-mediated excitatory postsynaptic currents (EPSCs) evoked by electrical stimulation. The NMDA/non-NMDA ratio of EPSC amplitude was high and the decay time course of NMDA-EPSC was slow in non-serotonergic/GABAergic neurons. In contrast, serotonergic neurons exhibited a low NMDA/non-NMDA ratio and a fast decay time course of NMDA-EPSC. Peripheral nerve ligation-induced chronic pain was associated with an increased NMDA/non-NMDA ratio in serotonergic neurons. Additionally, single-cell real-time RT-PCR analysis showed that peripheral nerve ligation up-regulated NR2B subunit expression in non-serotonergic/non-GABAergic neurons. Such changes in NMDA receptor expression in the PAG result in an alteration of the descending modulation of nociception, which might be an underlying mechanism for peripheral nerve injury-evoked persistent pain. Finally, the expression of NMDA receptors seems differentially regulated among neurons of different neurotransmitter phenotypes in the PAG.

Functional roles of endogenous d-serine in the chronic pain-induced plasticity of NMDAR-mediated synaptic transmission in the central amygdala of mice

The amygdala is implicated in chronic pain-induced emotional changes. Chronic pain induces plastic changes of the N-methyl-d-aspartate receptor (NMDAR) functions in the brain including the amygdala. d-Serine is synthesized endogenously by serine racemase and modulates NMDAR-mediated synaptic transmission as a coagonist of glycine binding site. To clarify the functional roles of endogenous d-serine in chronic pain-induced plasticity of NMDAR mediated synaptic transmission, we investigated the NMDAR-mediated excitatory synaptic current (EPSC) of neurons in the latero-capsular division of the central amygdala (CeLC) using brain slices from serine racemase knockout (SR-KO) mice with chronic pain induced by monoarthritis. The decay time of NMDAR-mediated EPSC was significantly elongated by monoarthritis in wild type (WT) mice, but not in SR-KO mice. The d-serine application-induced increase of NMDAR-mediated EPSC was significantly facilitated by monoarthritis in WT mice, but not in SR-KO mice. These results suggest that endogenous d-serine facilitates chronic pain-induced plastic changes of NMDAR mediated synaptic transmission in CeLC.

Differential expression of NMDA receptor subunits between neurons containing and not containing enkephalin in the mouse embryo spinal cord

We transfected cultures of mouse spinal cord slices with the enhanced green fluorescent protein (GFP) gene driven by the promoter for preproenkephalin, using the particle-mediated gene transfer system adapted for small neurons in the superficial dorsal horn, and observations were made after 4-6 days in vitro. A considerable number of cells in the superficial dorsal horn were observed to express GFP fluorescence, reminiscent of the previously reported distribution of enkephalinergic neurons in the spinal cord. The number of GFP-expressing neurons increased in response to forskolin application. Reverse transcription-polymerase chain reaction (RT-PCR) analysis of single neurons revealed that the N-methyl-d-aspartate (NMDA) receptor NR2B subunit is expressed more frequently in enkephalinergic neurons, and the NR2A subunit more frequently in non-enkephalinergic neurons. These observations suggest that expression of NMDA receptor subunits is controlled differentially in distinct populations of neurochemically identified neurons in the spinal cord. Biolistic particle-mediated gene transfection seems useful for identifying neuronal phenotypes in organotypic cultures of the spinal cord.

Effects of pregabalin on spinal d-serine content and NMDA receptor-mediated synaptic transmission in mice with neuropathic pain

Pregabalin (PGB) is a chemical derivative of the inhibitory neurotransmitter γ-aminobutyric acid, and is successfully used for the treatment of neuropathic pain. Substantial evidence suggests that d-serine, an endogenous co-agonist at the strychnine-insensitive glycine site of the NMDA receptor, counteracts the antinociceptive actions of PGB at the level of the spinal cord. In the present study, we examined the impact of PGB treatment on spinal d-serine content and NMDA receptor-mediated synaptic transmission in the superficial dorsal horn of peripheral nerve-ligated neuropathic mice. Mechanical allodynia was assessed using von Frey filaments. On post-surgical day 9 (after 5days of treatment with PGB [50mg/kg] or saline vehicle), the lumbar spinal cord was removed, homogenized, and ultrafiltrated. Supernatant samples were treated with Marfeys reagent and analyzed with liquid chromatography-mass spectrometry to measure d-serine content. In the electrophysiological experiments, tight-seal whole-cell recording was performed on neurons in the superficial dorsal horn of spinal cord slices. Partial sciatic nerve ligation increased spinal d-serine content, increased the NMDA/non-NMDA ratio of EPSC amplitudes, and slowed the decay phase of the NMDA component of EPSCs (NMDA-EPSCs). PGB treatment attenuated mechanical allodynia and reduced spinal d-serine content, decreased the NMDA/non-NMDA ratio, and shortened the decay time of NMDA-EPSCs. Furthermore, bath-applied d-serine attenuated the effects of PGB treatment. Although the precise mechanism for the effect of PGB on d-serine metabolism and abundance is unknown, the antinociceptive action of PGB likely involves the reduction of spinal d-serine content and subsequent attenuation of NMDA receptor-mediated synaptic transmission in the superficial dorsal horn.

Functional roles of endogenous D-serine in pain-induced ultrasonic vocalization.

The N-methyl-D-aspartate receptor (NMDAR) is crucial for pain-related behaviors. D-Serine is synthesized from L-serine by serine racemase (SR) and modulates NMDAR functions by acting as an agonist at the glycine-binding site. We analyzed noxious stimulus-induced ultrasonic vocalization and locomotor activity in the open-field test using SR knockout (SR-KO) mice to examine the role of endogenous D-serine in mammalian behaviors. SR-KO mice emitted less ultrasonic vocalization after noxious stimulation (VAS) than wild-type (WT) mice. The locomotor activity of WT mice decreased with repeated daily exposures to the open field, whereas that of SR-KO mice remained unchanged. VAS was significantly enhanced during arthritis in WT mice, whereas it was not enhanced during arthritis in SR-KO mice. These results indicate that mice lacking the ability to produce D-serine endogenously in the brain differ from normal mice with respect to the chronic pain-induced behavioral changes.