An association study of monoamine oxidase A (MAOA) gene polymorphism in methamphetamine psychosis

Methamphetamine continues to be the most widely abused drug in Japan. Chronic methamphetamine users show psychiatric signs, including methamphetamine psychosis. Monoamine oxidase A (MAOA) is one of the major enzymes responsible for the degradation of neurotransmitters. Abnormalities in MAO levels have been related to a wide range of psychiatric disorders. We examined whether or not the MAOA-u variable-number tandem repeat (VNTR) has a functional polymorphism in methamphetamine psychosis and whether or not such a polymorphism is related to the prolongation of psychosis. As expected, there was a significant difference in the MAOA-u VNTR between males with persistent versus transient methamphetamine psychosis (p = 0.018, odds ratio (OR) = 2.76, 95% CI: 1.18–6.46). Our results suggest that the high-activity allele class of MAOA-u VNTR in males may be involved in susceptibility to a persistent course of methamphetamine psychosis. We found no differences among females. The sample size of females with methamphetamine psychosis was too small to have significant analysis.     

Preferential suppression of limbic Fos expression by intermittent hypoxia in obese diabetic mice

Sleep apnea (SA) causes not only sleep disturbances, but also neurocognitive impairments and/or psychoemotional disorders. Here, we studied the effects of intermittent hypoxia (IH) on forebrain Fos expression using obese diabetic db/db mice to explore the pathophysiological alterations in neural activities and the brain regions related to SA syndrome. Male db/db mice were exposed to IH stimuli (repetitive 6-min cycles of 1 min with 5% oxygen followed by 5 min with 21% oxygen) for 8 h (80 cycles) per day or normoxic condition (control group) for 14 days. Fos protein expression was immunohistochemically examined a day after the last IH exposure. Mapping analysis revealed a significant reduction of Fos expression by IH in limbic and paralimbic structures, including the cingulate and piriform cortices, the core part of the nucleus accumbens and most parts of the amygdala (i.e., the basolateral and basomedial amygdaloid nuclei, cortical amygdaloid area and medial amygdaloid nucleus). In the brain stem regions, Fos expression was region-specifically reduced in the ventral tegmental area while other regions including the striatum, thalamus and hypothalamus, were relatively resistant against IH. In addition, db/db mice exposed to IH showed a trend of sedative and/or depressive behavioral signs in the open field and forced swim tests. The present results illustrate that SA in the obese diabetic model causes neural suppression preferentially in the limbic and paralimbic regions, which may be related to the neuropsychological disturbances associated with SA.     

Bilateral lesions of the central but not anterior or posterior parts of the piriform cortex retard amygdala kindling in rats

The piriform cortex is thought to be involved in temporal lobe seizure propagation, such as that occurring during kindling of the amygdala or hippocampus. A number of observations suggested that the circuits of the piriform cortex might act as a critical pathway for limbic seizure discharges to assess motor systems, but direct evidence for this suggestion is scarce. Furthermore, the piriform cortex is not a homogeneous structure, which complicates studies on its role in limbic epileptogenesis. We have previously reported data indicating that the central part of the piriform cortex might be particularly involved during amygdala kindling. In order to further evaluate the role of different parts of the piriform cortex during kindling development, we bilaterally destroyed either the central, anterior or posterior piriform cortex by microinjections of ibotenate two weeks before onset of amygdala kindling. Lesions of the anterior piriform cortex hardly affected kindling acquisition, except that fewer animals exhibited stage 3 (unilateral forelimb) seizures compared to sham controls. Lesions of the central piriform cortex significantly retarded kindling, which was due to a decreased progression from stage 3 to stage 4/5 seizures, i.e. the lesioned rats needed significantly longer for the acquisition of generalized clonic seizures in the late stages of kindling development. Lesions of the posterior piriform cortex did not significantly affect kindling development.The data demonstrate that different parts of the piriform cortex mediate qualitatively different effects on amygdala kindling. The central piriform cortex seems to be a neural substrate involved in the continuous development of kindling from stage 3 to stages 4/5, indicating that this part of the piriform cortex may have preferred access, either directly or indirectly, to structures capable of supporting generalized kindled seizure expression.     

Olfactory system modulation of hippocampal cell death

The hippocampal dentate gyrus is a major recipient of olfactory input in rodents, via connections from the olfactory (piriform) cortex and the olfactory bulb to the entorhinal cortex. Given this connectivity and the known role of activity in dentate gyrus granule cell survival, the present experiment examined the immediate effects of loss of olfactory input to the hippocampus on apoptosis. Adults rats underwent unilateral or bilateral olfactory bulb ablations (OBX), and allowed to recover 24–72 h before the piriform cortex and hippocampal dentate gyrus were processed for terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling [TUNEL] of apoptotic cells. OBX transiently increased TUNEL-positive cells in the ipsilateral piriform cortex and dentate gyrus. Increased TUNEL-labeling was apparent within 24 h in both structures, but was more extensive and prolonged in piriform cortex. The results suggest a trans-synaptic regulation of cell survival through at least two synapses.     

State-dependent functional connectivity of rat olfactory system assessed by fMRI

Functional connectivity between the piriform cortex and limbic and neocortical areas was assessed using functional magnetic resonance imaging (fMRI) of urethane anesthetized rats that spontaneously cycled between slow-wave and fast-wave states. Slow-wave and fast-wave states were determined indirectly through monitoring of respiration rate, which was confirmed to co-vary with state as determined by electrophysiological recordings. Previous electrophysiological data have suggested that the piriform cortex shifts between responsiveness to afferent odor input during fast-wave states and enhanced functional connectivity with limbic areas during slow-wave state. The present results demonstrate that fMRI-based resting state functional connectivity between the piriform cortex and both limbic and neocortical areas is enhanced during slow-wave state compared to fast-wave state using respiration as an indirect measure of state in urethane anesthetized rats. This state-dependent shift in functional connectivity may be important for sleep-dependent odor memory consolidation.     

Spontaneous oscillatory burst activity in the piriform-amygdala region and its relation to in vitro respiratory activity in newborn rats

The amygdala is important for the formation of emotions that are affected by olfactory information. The piriform cortex is involved in information processing related to olfaction. To investigate functional interactions between the piriform cortex and amygdala and their relation to medullary respiratory activity, we developed a novel in vitro preparation including the limbic system, brainstem, and spinal cord of newborn rats. With the use of optical and electrophysiologic recordings, we analyzed spontaneous neuronal activity in the piriform-amygdala complex in limbic-brainstem-spinal cord preparations from 0- to 1-day-old rats. For optical recordings, the preparation was stained with a voltage-sensitive dye, and inspiratory activity was monitored from the fourth cervical (C4) ventral root. Spontaneous oscillatory burst activity (up to 10/min) was detected from the rostral cut surface of limbic and para-limbic regions including the piriform cortex and amygdala. The burst activity initially appeared in the piriform cortex and then propagated to the amygdala. We averaged the imaging data in the limbic area with the use of C4 inspiratory activity as a trigger signal. The results suggest functional coupling of the rhythmic burst activity in the piriform-amygdala complex to medullary inspiratory activity, which was confirmed electrophysiologically by cross-correlation analysis of these signals. This rhythmic burst activity may be involved in the development of neuronal circuits that process information related to olfaction, emotion, and respiration.     

c－fos基因在听源性惊厥易感大鼠脑的表达──原位杂交组织化学法研究

运用原位杂交技术，以遗传性听源性惊厥易感大鼠Ｐ７７ＰＭＣ为对象，发现听源性惊厥可诱导大鼠脑内ｃ－ｆｏｓ基因快速、大量、短暂性表达。ｃ－ｆｏｓｍＲＮＡ分布于大脑皮层、梨状皮层、杏仁复合体、海马齿状回、上丘脑、背侧丘脑、下丘脑部分核团、下丘、蜗神经核、蓝斑及小脑等处。惊厥后皮层下结构中ｃ－ｆｏｓ基因表达变化程度超过皮层的变化，尤其是下丘、蜗神经核与惊厥时程有明显关系。推测皮层下结构对听源性惊厥的发生有重要意义。Ｐ＜０．０１讨论本文结果说明听源性惊厥同其它因素诱导的惊厥一样［３］，可诱导大鼠脑内ｃ－ｆｏｓ基因的表达，表达涉及到大脑皮层、海马齿状回、丘脑、下丘、蜗神经核等结构，其中以皮层下结构如丘脑、下丘、蜗神经核表达变化最显著。原位杂交显示的ｃ－ｆｏｓ基因表达特征类似于Ｎｏｒｔｈｅｒｎ杂交结果即快速、大量和短暂性。由于不同部位在惊厥活动中的作用差别，因此用原位杂交可以显示每一结构内ｃ－ｆｏｓ基因表达特点。如在惊厥后３０ｍｉｎ，海马齿状回中７０％以上的神经元单位胞质面积上银粒计数超过２０个，而梨状皮层及运动皮层仅占５～１３．８％。有报告指出海马齿状回为钙离子通道和ＮＭＤＡ受体高密度区域［４］，推测Ｃａ２＋和ＮＭＤＡ?     

Ectopic galanin expression and normal galanin receptor 2 and galanin receptor 3 mRNA levels in the forebrain of galanin transgenic mice

The functional interactions of the neuropeptide galanin (GAL) occur through its binding to three G protein-coupled receptor subtypes: galanin receptor (GALR) 1, GALR2 and GALR3. Previously, we demonstrated that GALR1 mRNA expression was increased in the CA1 region of the hippocampus and discrete hypothalamic nuclei in galanin transgenic (GAL-tg) mice. This observation suggested a compensatory adjustment in cognate receptors in the face of chronic GAL exposure. To evaluate the molecular alterations to GALR2 and GALR3 in the forebrain of GAL overexpressing mice, we performed complementary quantitative, real-time PCR (qPCR), in situ hybridization, and immunohistochemistry in select forebrain regions of GAL-tg mice to characterize the neuronal distribution and magnitude of GAL mRNA and peptide expression and the consequences of genetically manipulating the neuropeptide GAL on the expression of GALR2 and GALR3 receptors. We found that GAL-tg mice displayed dramatic increases in GAL mRNA and peptide in the frontal cortex, posterior cortex, hippocampus, septal diagonal band complex, amygdala, piriform cortex, and olfactory bulb. Moreover, there was evidence for ectopic neuronal GAL expression in forebrain limbic regions that mediate cognitive and affective behaviors, including the piriform and entorhinal cortex and amygdala. Interestingly, regional qPCR analysis failed to reveal any changes in GALR2 or GALR3 expression in the GAL-tg mice, suggesting that, contrary to GALR1, these receptor genes are not under ligand-mediated regulatory control. The GAL-tg mouse model may provide a useful tool for the investigation of GAL ligand-receptor relationships and their role in normal cognitive and affective functions as well as in the onset of neurological disease.     

Inhibiting dopamine reuptake blocks the induction of long-term potentiation and depression in the lateral entorhinal cortex of awake rats

Synaptic plasticity in olfactory inputs to the lateral entorhinal cortex may result in lasting changes in the processing of olfactory stimuli. Changes in dopaminergic tone can have strong effects on basal evoked synaptic responses in the superficial layers of the entorhinal cortex, and the current study investigated whether dopamine may modulate the induction of long-term potentiation (LTP) and depression (LTD) in piriform cortex inputs to layer II of the lateral entorhinal cortex in awake rats. Groups of animals were pretreated with either saline or the selective dopamine reuptake inhibitor GBR12909 prior to low or high frequency stimulation to induce LTD or LTP. In saline-treated groups, synaptic responses were potentiated to 122.4 ± 6.4% of baseline levels following LTP induction, and were reduced to 84.5 ± 4.9% following induction of LTD. Changes in synaptic responses were maintained for up to 60 min and returned to baseline levels within 24 h. In contrast, induction of both LTP and LTD was blocked in rats pretreated with GBR12909. Dopaminergic suppression of synaptic plasticity in the entorhinal cortex may serve to restrain activity-dependent plasticity during reward-relevant behavioral states or during processing of novel stimuli.     

Dysfunctions within limbic–motor networks in amyotrophic lateral sclerosis

Previous studies have shown that affective symptoms are part of the clinical picture in amyotrophic lateral sclerosis (ALS), the most common motor neuron disorder in elderly people. Diffuse neurodegeneration of limbic regions (e.g., prefrontal cortex [PFC], amygdala) was demonstrated in ALS post-mortem, although the mechanisms of emotional dysregulation in ALS in vivo remain unclear. Using functional imaging, we assessed the brain responses to emotional faces in 11 cognitively unimpaired ALS patients and 12 healthy controls (HCs). We tested whether regional activities and connectivity patterns in the limbic system differed between ALS patients and HCs and whether the variability in clinical measures modulated the neuroimaging data. Relative to HCs, ALS patients displayed greater activation in a series of PFC areas and altered left amygdala–PFC connectivity. Anxiety modulated the right amygdala–PFC connectivity in HCs but not in ALS patients. Reduced right premotor cortex activity and altered left amygdala–supplementary motor area connectivity were associated with longer disease duration and greater disease severity, respectively. Our findings demonstrate dysfunctions of the limbic system in ALS patients at early stages of the disease, and extend our knowledge about the interplay between emotional brain areas and the regions traditionally implicated in motor control.     

Bilateral microinjections of vigabatrin in the central piriform cortex retard amygdala kindling in rats

The piriform cortex (PC) is the largest region of the mammalian olfactory cortex with strong connections to limbic structures, including the amygdala, hippocampus, and entorhinal cortex. Various previous studies in rodents suggest that the PC might be very important in the development and maintenance of limbic kindling, i.e. a widely used model of temporal lobe epilepsy. GABAergic inhibition in the transition zone between the anterior and posterior PC, termed here central PC, seems to be particularly involved in the processes leading to progression of kindled seizures. This prompted us to study whether elevation of GABA levels in this subregion of the PC by bilateral microinjection of vigabatrin is capable of suppressing amygdala kindling. Rats were stimulated once daily until fully kindled (stage 5) seizures had developed. Vigabatrin (10 microg) was injected 24 h before the first stimulation as well as 6 h before the 5th and 10th stimulation, which approximately doubled the number of stimulations required for kindling development compared with controls. This marked retardation of kindling acquisition was predominantly due to a significant inhibition of the progression from stage 1 to stage 2 and stage 3 to stage 4 seizures, demonstrating that microinjection of vigabatrin into the central PC markedly inhibits the progression and secondary generalization of focal seizures emanating from the amygdala.     

Response inhibition deficits in mice with septal, amygdala, or cingulate cortical lesions

Mice with lesions of the septum, cingulate cortex, lateral amygdala or with combined lateral amygdala and septal lesions showed significant deficits in suppressing shock contingent activity. Animals with combined septal-lateral amygdala lesions showed the greatest deficits; animals with cingulate cortical damage showed only minimal deficits. No deficits were found in mice with medial amygdala lesions. The results are in general agreement with previous studies and provide additional evidence for the role of the limbic system in modulating response inhibition.     

Differential brain response to alcohol cue distractors across stages of alcohol dependence

Altered attention to alcohol-related cues is implicated in the craving and relapse cycle characteristic of alcohol dependence (ALC). Prior cue reactivity studies typically invoke explicit attention to alcohol cues, so the neural response underlying incidental cue exposure remains unclear. Here, we embed infrequent, task-irrelevant alcohol and non-alcohol cues in an attention-demanding task, enabling evaluation of brain responses to distracting alcohol cues. Alcohol dependent individuals, across illness phase (n = 44), and controls (n = 20) performed a cue-reactivity fMRI target detection task. Significant Group-by-Distractor effects were observed in dorsal anterior cingulate cortex (ACC), inferior parietal lobule, and amygdala. Controls and long-term abstainers increased recruitment of attention and cognitive control regions, while recent and long-term abstainers decreased limbic recruitment to alcohol distractors. Across phases of ALC, self-reported craving positively correlated with cue-related activations in ventral ACC, medial prefrontal cortex, and cerebellum. Results indicate that brain responses elicited by incidental alcohol cues differentiate phases of ALC.     

Repeated methamphetamine treatment increases expression of TRPV1 mRNA in the frontal cortex but not in the striatum or hippocampus of mice

The transient receptor potential vanilloid type 1 (TRPV1) is a non-selective ligand-gated cationic channel. The distribution of TRPV1 mRNA in various regions of the brain has been successfully established. Methamphetamine (MAP) is a psychostimulant and a major drug of abuse in many parts of the world. The powerful rewarding properties of MAP are attributed to multiple pharmacological actions, but the mechanistic association between TRPV1 expression and MAP-induced drug addiction has not established. In the present study, we conducted a time-course analysis of TRPV1 mRNA levels in the frontal cortex, striatum, and hippocampus of mouse brain following repeated MAP (2 mg/kg, i.p.) treatment. Our results demonstrate that expression of TRPV1 mRNA is significantly increased 1, 2, 6, 24, 48 h, and 1 week after the last MAP treatment in the frontal cortex but not in the striatum or hippocampus. These data support a potential role for TRPV1 in the treatment of MAP-induced drug addiction.     

A sensitizing d-amphetamine dose regimen induces long-lasting spinophilin and VGLUT1 protein upregulation in the rat diencephalon

Numerous studies in this lab and others have reported psychostimulant-induced alterations in both synaptic protein expression and synaptic density in striatum and prefrontal cortex. Recently we have shown that chronic d-amphetamine (d-AMPH) administration in rats increased synaptic protein expression in striatum and limbic brain regions including hippocampus, amygdala, septum, and paraventricular nucleus of the thalamus (PVT). Potential synaptic changes in thalamic nuclei are interesting since the thalamus serves as a gateway to cerebral cortex and a nodal point for basal ganglia influences. Therefore we sought to examine drug-induced differences in synaptic protein expression throughout the diencephalon. Rats received an escalating (1–8 mg/kg) dosing regimen of d-AMPH for five weeks and were euthanized 28 days later. Radioimmunocytochemistry (RICC) revealed significant upregulation of both spinophilin and the vesicular glutamate transporter, VGLUT1, in PVT, mediodorsal (MD), and ventromedial (VM) thalamic nuclei as well as in lateral hypothalamus (LH) and habenula. Strong positive correlations were observed between VGLUT1 and spinophilin expression in PVT, medial habenula, MD, VM and LH of d-AMPH-treated rats. No significant d-AMPH effect was seen in sensorimotor cortices for either protein. Additionally, no significant differences in the general vesicular protein synaptophysin were observed for any brain region. These findings add to evidence suggesting that long-lasting stimulant-induced synaptic alterations are widespread but not ubiquitous. Moreover, they suggest that d-AMPH-induced synaptic changes may occur preferentially in excitatory synapses.     

Distribution of HSP72 induction and neuronal death following limbic seizures.

A small area of deep prepiriform cortex is uniquely susceptible to convulsant and anticonvulsant drugs in the rat. We have studied the pattern of expression of the non-constitutive stress protein (HSP72) following seizures induced by unilateral microinjection of bicuculline into this area. HSP was seen first in ipsilateral dorsal medial thalamus, amygdala and associated piriform cortex, and with more sustained seizures was seen bilaterally in these structures as well as in other projection sites. Neuronal cell death, as assessed by acid-fuchsin staining, occurred in the same brain regions. Frank necrosis was found in the ipsilateral piriform cortex with prolonged seizures. Behaviorally, the seizures induced are characteristic of involvement of the limbic system and, therefore, may be a model of human complex partial seizures.     

芳香化酶mRNA在小鼠脑内的表达及其分布

目的 研究芳香化酶ｍ（ＲＮＡ（ａｒｏｍａｔａｓｅ ｍＲＮＡ）在小鼠脑内的基因表达。方法 原位杂交组织化学和ＰＮＡ斑点杂交。结果 （１）斑点杂交结果显示,脑内芳香化酶ｍＲＮＡ在小鼠Ｅ１６－Ｐ３００整个发育过程中均有表达,表达高峰在生后６ｄ左右,成年后降至最低;（２）脑内芳香化酶ｍＲＮＡ主要定位于神经元;（３）芳香化酶ｍＲＮＡ在脑内的表达,阳性区域主要分布于大脑皮层,丘脑、下丘脑及边缘系统。其中,皮质锥体细胞层、内侧视前区、隔内侧核、海马各区锥体层、杏仁核群、扣带皮质、梨状前皮质及杏仁周皮质等部位阳性信号较强;中等强度的阳性信号见于丘脑腹内、外侧核,丘脑外侧背核、下丘脑室旁核、室周核等处。结论 以上结果进一步证明脑内芳香化酶的表达与脑发育存在一定的相关性,芳香化酶ｍＲＮＡ的表达部位与文献报道酶的活性分布基本一致;海     

Effects of corticotropin releasing factor on spontaneous burst activity in the piriform-amygdala complex of in vitro brain preparations from newborn rats

The amygdala is an important higher regulatory center of the autonomic nervous system, involved in respiratory and cardiovascular control, and it also plays a role in the formation of emotions. Corticotropin-releasing factor (CRF) is a neuropeptide involved in stress responses. We have examined the effects of CRF on the spontaneous burst activity in the piriform-amygdala complex of rat brain preparations in vitro. Limbic-brainstem-spinal cord preparations of 0- to 1-day-old Wistar rats were isolated under deep ether anesthesia, and were superperfused in a modified Krebs solution. Bath application of 50 nM CRF substantially increased the frequency of burst activity in the piriform-amygdala complex, whereas this polypeptide exerted only minor effects on C4 inspiratory activity. The excitatory effect of CRF on the amygdala burst was effectively blocked by the CRF1 antagonist, antalarmin, but not the CRF2 antagonist, astressin-2B, suggesting that CRF1 mediated the excitatory effect. The spatio-temporal pattern of the burst activity according to optical recordings was basically identical to the controls; the burst activity initially appeared in the piriform cortex and then propagated to the amygdala. The present experimental model could be useful for the study of role of the limbic system, including the amygdala, in stress responses.     

Age-related impairments in memory and in CREB and pCREB expression in hippocampus and amygdala following inhibitory avoidance training

This experiment examined whether age-related changes in CREB and pCREB contribute to the rapid forgetting seen in aged animals. Young (3-month-old) and aged (24-month-old) Fischer-344 rats received inhibitory avoidance training with a low (0.2 mA, 0.4 s) or moderate (0.5 mA, 0.5 s) foot shock; memory was measured 7 days later. Other rats were euthanized 30 min after training, and CREB and pCREB expression levels were examined in the hippocampus, amygdala, and piriform cortex using immunohistochemistry. CREB levels decreased with age in the hippocampus and amygdala. After training with either shock level, young rats exhibited good memory and increases in pCREB levels in the hippocampus and amygdala. Aged rats exhibited good memory for the moderate but not the low shock but did not show increases in pCREB levels after either shock intensity. These results suggest that decreases in total CREB and in pCREB activation in the hippocampus and amygdala may contribute to rapid forgetting in aged rats. After moderate foot shock, the stable memory in old rats together with absence of CREB activation suggests either that CREB was phosphorylated in a spatiotemporal pattern other than analyzed here or that the stronger training conditions engaged alternate mechanisms that promote long-lasting memory.