Chronic morphine treatment increases the expression of the neural cell adhesion molecule in the dorsal horn of the mouse spinal cord

It is well known that prolonged exposure to morphine results in tolerance to morphine-induced antinociception. In the present study, we found that mice that were tolerant to morphine-induced antinociception exhibited an increase in immunoreactivity for the neural cell adhesion molecule in the dorsal horn of the spinal cord, which was highly overlapped with immunoreactivity for the increased metabotropic glutamate receptor 5 induced by morphine. These findings support the idea that repeated stimulation of μ-opioid receptors increases the expression of neural cell adhesion molecule and metabotropic glutamate receptor 5. This phenomenon leads to the enhanced excitatory synaptic transmission in the dorsal horn of the spinal cord, and in turn suppresses the morphine-induced antinociception.     

Caudal end of the rat spinal dorsal horn

We have previously demonstrated that the transformation of the caudal spinal cord through the conus medullaris to the filum terminale takes place in three steps. In the conus medullaris the twin layers of CGRP-immunoreactive and IB4-labeled primary afferent fibers as well as the translucent portion of the superficial dorsal horn equivalent to the substantia gelatinosa discontinue before the complete removal of the dorsal horn. Parallel with these changes VGLUT1-immunoreactive myelinated primary afferent fibers arborize not only in the deep layers but also in the entire extension of the remaining dorsal horn, while scattered CGRP fibers still remains at the margin of and deep in the dorsal horn. PKCgamma-immunoreactive dorsal horn neurons discontinue parallel with the disappearance of the IB4-labeled nerve fibers. These observations suggest that in the dorsal horn certain neurons are linked to the substantia gelatinosa, while others are substantia gelatinosa-independent neurons.     

An investigation of depotentiation of long-term potentiation in the CA1 region of the hippocampus

We have investigated long-term synaptic depression in the CA1 region of rat hippocampal slices. Prolonged low-frequency stimulation (LFS; 900 stimuli delivered at 2 Hz) of the Schaffer collateral-commissural pathway in naïve slices did not induce long-term depression (LTD) of synaptic transmission. However, if long-term potentiation (LTP) was firstly induced in the pathway then LFS generated an LTD-like effect (i.e. depotentiation of LTP). Depotentiation could be induced 2 h (the longest time studied) after the induction of LTP and was stable for the duration of the experiment (followed for up to 40 min). The induction of depotentiation was not blocked by the N-methyl-d-aspartate receptor antagonist d-2-amino-5-phosphonopentanoate, the L-type voltage-gated Ca²⁺ channel blocker nimodipine or the nitric oxide synthase inhibitor N-nitro-l-arginine. However, the magnitude of depotentiation was reversibly reduced, in a stereoselective manner, by the specific metabotropic glutamate receptor (mGluR) antagonist (+)--methyl-4-carboxyphenylglycine. These results show that prolonged low frequency stimulation can result in an mGluR-dependent depotentiation of LTP.     

Evidence that Ca/calmodulin-dependent protein kinase mediates the modulation of the Ca2+-dependent K+ current,I AHP,by acetylcholine,but not by glutamate,in hippocampal neurons

Muscarinic and metabotropic glutamate receptor agonists increase the excitability of hippocampal and other cortical neurons by suppressing the Ca²⁺-activated K⁺current,I _AHP, which underlies the slow afterhyperpolarization (AHP) and spike frequency adaptation. We have examined the mechanism of action of a muscarinic agonist (carbachol) and a metabotropic glutamate receptor agonist (1-Aminocyclopentane-trans-1,3-dicarboxylic acid; t-ACPD) onI _AHP in hippocampal CA1 neurons in slices, by using highly specific protein kinase inhibitors. We found that inhibition of protein kinase A (PKA) with the adenosine 3,5-cyclic monophosphate (cAMP) analogue Rp-adenosine-3,5-cyclic phosphorothioate Rp-cAMPS, did not prevent the muscarinic and glutamatergic suppression ofI _AHP. In contrast, two specific peptide inhibitors of Ca²⁺/calmodulin-dependent protein kinase II (CaM-K II), each partially blocked the effect of carbachol, but not the effect of t-ACPD onI _AHP. We conclude that CaM-K II, but not PKA, is involved in mediating the muscarinic suppression ofI _AHP, although other pathways may also contribute. In contrast, neither CaM-K II nor PKA seems to mediate the metabotropic glutamate receptor action onI _AHP.     

Activation of metabotropic glutamate receptor 3 enhances interleukin (IL)-1beta-stimulated release of IL-6 in cultured human astrocytes

Previous studies have demonstrated that human astrocytes express mRNA and receptor protein for group I and II metabotropic glutamate receptors (mGluRs). Whether these receptors can influence the inflammatory and immune response and can modulate the capacity of astrocytes to produce inflammatory cytokines is still unclear. Inflammatory cytokines can be produced by activated glial cells and play a critical role in several neurological disorders. Astrocyte-enriched human cell cultures growing in a serum-free chemically defined medium were used to study the regulation of IL (interleukin)-1β and IL-6 in response to mGluR activation. Astrocytes cultured in the absence or in the presence of epidermal growth factor (EGF), did not secrete significant IL-1β and IL-6, as determined by specific enzyme-linked immunosorbent assay (ELISA). Activation of mGluRs using (S)-3,5-dihydroxyphenylglycine (DHPG; selective group I agonist) or DCG-IV (selective group II agonist) did not affect the production of interleukins under both growth conditions. On exposure to IL-1β high levels of IL-6 were detected. Activation of mGluR3 with DCG-IV (but not of mGluR5 with DHPG) enhanced, in the presence of IL-1β, the release of IL-6 in a dose dependent manner in astrocytes cultured under conditions (+EGF) in which the mGluR expression is known to be upregulated. The effect of mGluR3 activation on IL-1β stimulated release of IL-6 was prevented by selective group II mGluR antagonists. The capacity of mGluR3 to modulate the release of IL-6 in the presence of IL-1β supports the possible involvement of this receptor subtype in the regulation of the inflammatory and immune response under pathological conditions associated with glial cell activation.     

Growth,regression and cell death in rat liver as related to tissue levels of the hepatomitogen cyproterone acetate

Previous studies have shown that xenobiotic compounds such as the environmental pollutant -hexa-chlorocyclohexane (-HCH) and the synthetic sex steroid cyproterone acetate (CPA) induce growth of rat liver by hypertrophy and hyperplasia. After withdrawal of the growth stimuli, liver hypertrophy was usually found to be readily reversible. Conflicting observations were made concerning the fate of liver hyperplasia: hepatic hyperplasia persisted when induced by -HCH but was found to be partially reversible when induced by CPA. The present study confirms the reversibility of hepatic hyperplasia induced by CPA in rats: about 30% of liver DNA present at maximal liver enlargement disappeared within 6 days after cessation of CPA treatment. Simultaneously, a dramatic increase in the rate of cell elimination by apoptosis was found. Glutamate-pyruvate transaminase and alkaline phosphatase in serum did not show major increases, suggesting that cell death was not due to lytic membrane damage. Furthermore, if treatment with CPA was continued or resumed, the enhanced DNA content persisted and the number of apoptotic bodies was greatly reduced. These observations suggest that the occurrence of cell death is due to withdrawal of the growth stimulus CPA. It may reflect a regulatory phenomenon serving to maintain homeostasis of cell number.Further studies showed that CPA is rapidly eliminated from rat liver and serum: t 1/2 in the liver is about 11 h. In contrast, -HCH was previously found to be eliminated more slowly: t 1/2 approximately 144 h. The present study revealed that -HCH, CPA and nafenopin lower the number of apoptotic bodies. This suggests that inducers of liver growth can inhibit hepatocellular death by apoptosis. It is concluded that the regression of hyperplasia after CPA withdrawal may be due to its rapid elimination. On the other hand the relatively long persistence of -HCH may result in inhibition of cell death and thereby stabilize hepatic hyperplasia.Abbreviations CPA cyproterone acetate - -HCH -hexachlorocyclohexane - PB phenobarbital - NAF nafenopin - AB apoptotic body - b.w. body weight - p. admin. post-administration - GPT glutamate-pyruvate transaminase - ALP alkaline phosphatase Dedicated to Professor W. Koransky on the occasion of his 65th birthday     

一氧化氮对谷氨酸单钠脑损害小鼠学习记忆能力的影响

目的　观察谷氨酸单钠 (MSG)对小鼠学习记忆的影响及血浆、脑内NO含量的变化。方法　给断乳分窝小鼠MSG灌胃 ,每天 2次 ,连续 30d ,31d早灌胃后对小鼠进行迷宫行为训练 ,2 4h后对其进行迷宫记忆检测 ,用硝酸还原酶法检测血浆脑内的NO含量。结果　MSG对小鼠学习记忆能力有影响并存在剂量效应关系 ,其血浆、脑中NO含量均升高并有显著差异 (P <0 .0 5 )。结论　NO可加重MSG对小鼠学习与记忆的损害作用。     

Plasticity of GABA- and glutamate-containing terminals in the mouse thalamic ventrobasal complex deprived of vibrissal afferents: an immunogold-electron microscopic study

GABA and glutamate immunogold staining demonstrated that nerve cells of the thalamic ventrobasal complex (VB) of mice were positive exclusively for glutamate. None of the neuronal perikarya reacted the GABA antibody. By using alternate thin sections of the normal VB, it was also shown that large "specific" somatosensory and small corticothalamic terminals, both of which contained spherical synaptic vesicles, exhibited only glutamate-like immunoreactivity. A third axonal type, containing flat-ovoid synaptic vesicles, stained only for GABA. Seventy-five days after coagulation of the vibrissal follicles in newborn mice, a characteristic multiplication of GABA positive axon terminals was observed. In addition, it was demonstrated that, similarly to modified cortical endings (Hámori et al., J. Comp. Neurol. 254:166-183, '86), many GABA positive terminals appeared as specific afferent endings, replacing the missing "specific" vibrissal afferents. This finding shows a remarkable plasticity of inhibitory GABA axons during developmental synaptogenesis and provides further evidence that the size, location, and the type of attachment of presynaptic terminals are dependent on their postsynaptic target.     

Cross-desensitization Reveals Pharmacological Specificity of Excitatory Amino Acid Receptors in Isolated Hippocampal Neurons

Ionic currents elicited by excitatory amino acids were studied, using the concentration clamp method, in enzymatically isolated rat hippocampal neurons. Cross-desensitization between the responses to various agonists was applied to separate the activity of two types of receptors, N-methyl-d-aspartate (NMDA) and non-NMDA. NMDA receptors were selectively activated by NMDA, l- and d-aspartate, d-glutamate and quinolinate. Kainate and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate appeared to be selective, and quisqualate relatively less selective non-NMDA agonists, acting on the same receptor type. l-Glutamate, l- and d-homocysteate activated both receptor types. It is supposed that two receptor sites, activation site and desensitization site, control the action of agonists at the non-NMDA receptor. When examined in the cross-desensitization experiments, NMDA and non-NMDA receptors appear to be represented by the two homogeneous and independent receptor populations operating different ionic channels.     

Regional alterations of brain biogenic amines and GABA/glutamate levels in rats following chronic lead exposure during neonatal development

Wistar rat pups were administered either a high dose of lead acetate (400 g lead/g body weight/day) or a low dose (100 g lead/g body weight/day) by gastric intubation, from 2 days through 60 days of age. The rats on both these doses exhibited statistically significant decreases in body and brain weights throughout the lead treatment period. A group of rats on high dose was also rehabilitated by discontinuing the lead from 60 days of age. In these rats, at 160 days of age, the body weight but not the brain weight recovered to normal levels. During the lead intake, the rats on high dose revealed significant elevations in the levels of noradrenaline (NA) in the hippocampus (HI), cerebellum (CE), hypothalamus (HY), brainstem (BS), and accumbens-striatum (SA). The elevated levels in all the above regions except in the HY persisted even after rehabilitation. The dopamine (DA) levels changed significantly in opposite directions in HY (elevation) and BS (reduction) during the lead treatment, and the HY recovered after rehabilitation. Under lead, the serotonin (5HT) levels were elevated significantly in the HI, BS and MC (motor cortex), while after rehabilitation the abnormality persisted only in the MC. Low dose lead treatment was also effective on the same areas of brain. In the low dose group, estimation of the levels of GABA and glutamate were also done, and a significant decrease of GABA in CE and glutamate in MC was observed. The differences observed in the neurotoxic effects (none or significant) of lead in the different regions for each of the transmitters (NA, DA, 5HT) supports the interesting conclusion that the vulnerability of the axon terminals of any given type is dependent on some regional factors, although the projections of the different regions originate from an apparently similar category of neurons in the brain stem.