Several GABAA receptor subunits are expressed in LHRH neurons of juvenile female rats

Gamma aminobutyric acid (GABA), the dominant inhibitory neurotransmitter in brain, is involved in the developmental regulation of LHRH secretion. Morphological studies in rodents have demonstrated that LHRH neurons are innervated by GABA-containing processes, suggesting that LHRH secretion is under direct transsynaptic GABAergic control. While GABA acts through two different receptors, GABA_A and GABA_B, to exert its effects, it appears that GABA_A receptors are able to mediate both inhibitory and stimulatory effects of GABA on LHRH neurons. GABA_A receptors are heterooligomeric ligand-gated anion channels that exhibit a diverse array of functional and pharmacological properties. This diversity is determined by the structural heterogeneity of the receptors, which are assembled from the combination of different classes of subunits with multiple isoforms. Although several studies have described the effect of GABA_A receptor stimulation on LHRH and/or gonadotropin release in prepubertal animals, nothing is known about the receptor subunits that may be expressed in LHRH neurons at this phase in development. Double immunohistofluorescence followed by confocal laser microscopy revealed that subsets of prepubertal LHRH neurons are endowed with ₁, ₂, β_2/3, and γ₂ GABA_A receptor subunits. Combined immunohistochemistry for LHRH neurons and in situ hybridization for GABA_A subunit mRNAs confirmed that the genes encoding the ₁, ₂, β₃ and γ₂ subunits, but not the γ₁ subunit, are expressed in LHRH neurons. Notwithstanding the relative insensitivity of these methods, both the immunohistochemical and hybridization histochemical approaches employed indicate that only a fraction of LHRH neurons are endowed with GABA_A receptors. This arrangement suggests that those LHRH neurons bearing the appropriate GABA_A receptors are responsible for either the entire secretory response to direct GABAergic inputs or for its initiation.     

Discriminative stimulus properties of ethanol in the rat: effects of neurosteroids and picrotoxin

Tetrahydrodeoxycorticosterone (5β-THDOC; 1.3–12.0 mg/kg), a neurosteroid enhancing the GABA_A receptor-associated chloride conductance, produced predominantly ethanol-appropriate responding (>80%) in rats trained to discriminate 1.0 g/kg ethanol from saline. However, neither picrotoxin (0.25–1.5 mg/kg), nor dehydroepiandrosterone sulfate (0.01–100.0 mg/kg), a neurosteroid acting as a GABA_A receptor antagonist, attenuated the stimulus effects of ethanol. These results indicate that: (1) at least certain neurosteroids may produce subjective states similar to these induced by ethanol; (2) blockade of the GABA_A receptor-associated channel does not eliminate the ethanol interoceptive cue in rats. ©1997 Elsevier Science B.V. All rights reserved.     

Advanced lipid peroxidation end-products in Alexander's disease

Rosenthal fibers (RF), intra-astrocytic hyaline inclusions, accumulate in various pathological conditions and are the histological hallmark of Alexander's disease. While the major protein components of RF have been identified, the factors accounting for their pathogenesis, accumulation, and insolubility are largely unknown. In this study, we immunohistochemically examined three cases of Alexander's disease using antibodies to a lysine-derived pyrrole modification arising from 4-hydroxy-2-nonenal, a highly cytotoxic reactive aldehyde produced by lipid peroxidation. In all the cases of Alexander's disease examined, strong immunolabeling of RF by the antibodies to 4-hydroxy-2-nonenal pyrrole adducts were noted. By contrast, age-matched control cases showed no immunoreactivity. These results indicate that modification of protein by lipid peroxidation adducts may play an important role in the formation of RF as well as in the pathogenesis of Alexander's disease. Furthermore, taken together with our previous data indicating advanced Maillard reaction end products in RF, it seems that post-translational modification of RF, initiated by oxidative stress, is critical for both the accumulation and the insolubility of RF, and therefore, by inference, in the pathogenesis of Alexander's disease.     

Rotatory seizures in juvenile myoclonic epilepsy

Rotatory seizures have been reported in association with focal intracranial lesions. This type of seizure was also described in patients with primary generalized epilepsies. To our knowledge, there is only one previous publication denoted an association between juvenile myoclonic epilepsy (JME) and rotatory seizures. We present two female patients with JME and rotatory seizures together. The onset of myoclonic jerks and generalized tonic clonic (GTC) seizures was in their midteens. Their interictal EEGs showed bilateral symmetric spike and polyspike wave discharges. The rotatory seizures of the patients started at age of 26 and 33 years, respectively. In one of the patients, turning to the left was followed by three or four complete turns, after then, she had GTC seizures. The other patient has turned to the right with only one or two turns and sometimes continued with GTC seizures. Neuroradiologic investigations including brain CT, MRI, and SPECT were performed. Response to valproate therapy of rotatory seizures was good. We believe that rotatory seizures are rarely seen in JME patients, and this causes false diagnosis which lead unsuitable drug choice.     

Challenges in Duchenne muscular dystrophy

The last seven years has witnessed an explosion in our understanding of the muscular dystrophies. In the early 1980s, prenatal diagnosis of Duchenne muscular dystrophy was developed. The cloning of the gene, in 1996, resulted in a better understanding of the disease process and led to the identification of a novel complex at the membrane. This information led to the cloning of other genes responsible for the autosomally inherited dystrophies. As we approach the millenium, the challenge is shifting to the development of therapy of these diseases. This review, in honour of Professor Alan Emery, explains how these advances have had an impact in the clinical management of pateints and the promise the progress holds for the future.     

Ibogaine acts at the nicotinic acetylcholine receptor to inhibit catecholamine release

In an effort to determine mechanisms of action of the putative anti-addictive agent ibogaine, we have measured its effects on catecholamine release in a model neuronal system, cultured bovine chromaffin cells. Various modes of stimulating catecholamine release were used including nicotinic ACh receptor activation, membrane depolarization with elevated K⁺ and Na⁺ channel activation with veratridine. In addition, because ibogaine has been reported to interact with kappa opioid receptors, we tested whether kappa receptor antagonists could reverse ibogaine's effects on catecholamine release. Ibogaine, at low concentration (<10 μM) was found to selectively inhibit nicotinic receptor-mediated catecholamine release, while having no significant effect on release evoked by either veratridine or membrane depolarization with elevated K⁺. The inhibitory actions of ibogaine and the kappa agonists were not reversed by preincubation with the opioid antagonists nor-binaltorphimine or naltrexone, suggesting that these inhibitory effects are not mediated by the kappa opioid receptor. The effects of low dose (10 μM) ibogaine were rapidly reversible, while the inhibitory effects of higher ibogaine doses persisted for at least 19 h following ibogaine washout. The results provide evidence for a mechanism of action ibogaine at the nicotinic ACh receptor. The results are consistent with a model in which the initial high transient brain concentrations (100 μM) of ibogaine act at multiple cellular sites and then have a selective action at the nicotinic ACh receptor cation channel following its metabolism to lower brain concentrations. The present findings are relevant to potential anti-addictive actions of ibogaine and to the development of drugs to combat nicotine addiction.     

Estrogen facilitates induction of long term potentiation in the hippocampus of awake rats

In order to test the hypothesis that circulating levels of estrogen modulate synaptic plasticity in the hippocampus, we have studied the induction of long term potentiation (LTP) in awake rats. Ovariectomized animals, chronically implanted with a recording electrode in the cell body layer of CA1 and a stimulating electrode in stratum radiatum, were used to record evoked field potentials (population spike (PS) and summed EPSP) daily for at least 4 days before injection of sesame oil or 100 μg of estradiol benzoate per kg b.w. (E2). Basal levels of response to single square pulses (0.01 ms pulse width) delivered at 0.05 Hz through the stimulating electrode were recorded daily for 2 days after injection. To induce LTP a high-frequency `theta pattern' stimulation was administered. Basal recordings at low-frequency stimulation did not change after injection. After high-frequency stimulation all (7/7) E2 injected animals showed LTP whereas only 1/6 oil injected controls did so; the mean increase in amplitude of the PS and slope of the EPSP after high-frequency stimulation were significantly greater in E2 treated rats. Input/output curves did not change significantly after E2 administration. These results show that at low-frequency stimulation, transynaptic responses of pyramidal neurones in CA1 are not affected by changes in levels of circulating estrogen, while synaptic plasticity — which is at the basis of proposed hebbian associative memory — is facilitated by estrogen treatment.     

Specificity of interleukin–1β-induced changes in monoamine concentrations in hypothalamic nuclei: blockade by interleukin-1 receptor antagonist

The purpose of this study was to examine specificity in the effects of interleukin-1β (IL-1β) on monoamines in various areas of the hypothalamus. Adult male rats were injected i.p. with saline or 2.5 or 5.0 μg of IL-1β or were pretreated with 500 μg of IL-1 receptor antagonist (IL-1ra) followed 5 min later by 5 μg of IL-1β. The paraventricular nucleus (PVN), arcuate nucleus (AN), median eminence (ME), and medial preoptic area (MPA) were microdissected and analyzed for neurotransmitter concentrations by high-performance liquid chromatography with electrochemical detection (HPLC-EC). In the PVN, IL treatment produced significant increases in the concentrations of norepinephrine (NE), dopamine (DA), DA metabolite dihydroxyphenylacetic acid (DOPAC), serotonin (5-HT), and its metabolite 5-hydroxyindoleacetic acid (5-HIAA). IL-1 treatment increased the concentrations of NE and DA in the AN but only of NE in the ME, and it was without any effect in the MPA. Pretreatment with IL-1ra completely blocked the IL-1 effects. It is concluded that IL-1 induces highly specific changes in monoamine metabolism in the hypothalamus, and the nature of these changes depends on specific hypothalamic nuclei.     

AMPA/kainate receptors modulate the survival in vitro of embryonic brainstem cells

This study aimed at analyzing the involvement of (RS)--amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate (AMPA/kainate) receptors in the survival of cultured rat embryonic brainstem cells, dissociated on embryonic day 14. The cell number was estimated after pharmacological manipulation of the receptors by exposure to agonists or antagonists. The developmental stage at the moment of drug application was critical for cell survival. We observed after 8 days in vitro a much stronger decrease in the number of gamma-enolase-positive cells when the cultures were treated for 3 days with the antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX) starting on the day of plating than when DNQX was added after 5 days in vitro. Conversely, exposure to the agonists (RS)-2-amino-3-(3-hydroxy-5-tri-fluoromethyl-4-isoxazolyl)-propionic acid (T-AMPA) or kainate for 3 days significantly reduced cell survival only when the treatment was initiated after 5 days in vitro. Survival of S-100-positive cells was not affected after exposure to either agonists or antagonists. Neither agonist nor antagonist treatment modified cell proliferation, as assessed by 5-bromo-2′-deoxyuridine (BrdU) staining, suggesting that the decrease in the number of gamma-enolase-positive cells is essentially due to cell death. If some of the processes we observed in vitro correspond to analogous events in vivo, then exposure to excitatory amino acid receptor agonists or antagonists at critical stages of embryogenesis may alter the development of the central nervous system.     

Postnatal development of interlaminar astroglial processes in the cerebral cortex of primates

Long astroglial processes traversing several cortical laminae appear to be characteristic of primate brains. Whether interlaminar processes develop as a modification of radial glia or are truly postnatal elements stemming from stellate astroglia, could be assessed by analyzing their early developmental stages. A survey of glial fibrillar acidic protein immunoreactive (GFAP-IR) astroglial interlaminar processes in the cerebral cortex of Ceboidea monkeys at various postnatal developmental ages, and in human cortical samples of a ten day and a seven year old child disclosed that such processes develop postnatally. At one month of age GFAP-IR interlaminar processes in monkeys were scarce and short in most frontal, parietal or occipital (striate) cortical areas, except for sulcal (principal and orbital sulci) and temporal cortical areas. Some processes were weakly positive for vimentin, and these were most abundant in ventral temporal cortical areas. At two months of age processes were present in all these areas, albeit in restricted patches and significantly shorter than in adults. The expression of this pattern was increased at seven months of age. At three years of age almost every area showed abundant processes and with lengths comparable to the adult Ceboidea individuals. In humans, at 10 days of age long interlaminar processes were readily apparent in a frontal cortex sample, becoming most apparent at the age of seven years although not reaching yet the adult characteristics as described previously. Conclusions : (1) GFAP-IR interlaminar processes develop postnatally, thus typifying a subtype of the classical stellate forms ; (2) they bear no obvious direct relationship with radial glia ; (3) their development is not contemporary among the various cortical regions. These long cellular processes represent an addition to those already described for other astroglial cell types in the adult mammalian brain (Golgi–Bergmann glia, tanicytes, Muller cells) .     

Early-onset facioscapulohumeral muscular dystrophy: two case reports

This report concerns two patients with facioscapulohumeral muscular dystrophy (FSHD) whose facial weakness began in infancy. In both patients, biopsied muscle histology showed mild myogenic changes accompanied by some regenerating and some small angular fibers, while endomysial inflammatory cellular infiltration was observed in Patient 1. The finding that our very young patients had muscle histopathological findings compatible with classical FSHD supports the previously expressed view that muscle histopathology is not related to either age or duration of the disease. Although Patient 2 was a sporadic case, both patients had the abnormal EcoRI DNA fragment detected by Southern blot analysis with probes p13E-11 and pFR-1, a finding compatible with FSHD. This indicates that gene analysis of sporadic cases must be as significant as that of familial cases. This report on patients with very early-onset and with common muscle histopathological and molecular genetic findings should contribute to widening the clinical spectrum of FSHD.     

Serotoninergic innervation of stapedial and tensor tympani motoneurons

Retrograde tracing and neurotransmitter immunohistochemistry were combined to determine whether serotonin neurons innervated stapedial and tensor tympani motoneurons. With high-power light microscopy, putative axo-somatic and axo-dendritic contacts were observed between serotonin-positive endings and both stapedial and tensor tympani motoneurons, indicating that serotonin neurons terminate on brainstem motoneurons innervating the middle-ear muscles. With this connection, the serotonin system may directly modulate middle-ear muscle activity.     

Temporal pattern of plasminogen activator activity in the developing chick cerebellum

Plasminogen activators are considered to be involved in several developmental events. The present work aims at characterizing the developmental pattern of expression of plasminogen activators in the chick cerebellum.Soluble fractions derived by ultracentrifugation from Triton X-100 treated membrane fractions were used for determination of the enzyme activity with a radial fibrinolytic assay. By using specific inhibitors and different anti-plasminogen activators antibodies it is shown that only one type of the enzyme, the urokinase-type plasminogen activator, is expressed during the cerebellum ontogeny. Our results show the existence of a bimodal pattern of enzyme activity with two peaks that temporally coincide with the processes of massive neuronal migration, neurite outgrowth and synapse formation and plasticity. It is proposed that plasminogen activator could play a role in these developmental events and that its pattern of variability is developmentally regulated.     

Source of extracellular brain adenosine during hypoxia in fetal sheep

Microdialysis was performed to determine whether hypoxia increases fetal brain adenosine (ADO) concentration through dephosphorylation of extracellular 5′-adenosine monophosphate (5-AMP). Hypoxia (fetal PaO₂ ≈ 14 Torr) increased fetal brain ADO levels two-fold when the probes were perfused with synthetic cerebrospinal fluid (CSF) containing inhibitors of the nucleoside transporter but not with this solution plus a blocker of ecto-5′-nucleotidase (AOPCP). The hypoxia-induced rise in fetal brain ADO concentrations depends critically upon the hydrolysis of extracellular 5′-AMP.     

Dissociation Between Semantic and Autobiographic Memory: A Case Report

We report a patient who showed a dissociation between impaired semantic memory and preserved autobiographic memory. M.N., a 56-year-old right-handed woman, developed a supra sellar meningioma and underwent multiple operations. Following Lineac radiation, necrosis occurred in the left temporal lobe. Magnetic Resonance Imaging revealed bilateral lesions in the temporal lobes and in the right basal frontal lobe.

The patient's intellectual deterioration and anterograde amnesia were mild. Language was preserved, but for a subtle anomia. Autobiographic memory remained intact, while semantic memory for public events, historical figures, cultural items, knowledge of low frequency words and technical terms related to her profession was severely impaired. We hypothesize that bilateral lesions of the anterior half of the middle region of the temporal lobe plays a crucial role in causing deficits in semantic memory.     

Age-dependent changes of presynaptic neuromodulation via A1-adenosine receptors in rat hippocampal slices

Thepresynaptic neuromodulation of stimulation-evoked release of [³H]-acetylcholine by endogenous adenosine, via A₁-adenosine receptors, was studied in superfused hippocampal slices taken from 4-, 12- and 24-month-old rats. 8-Cyclopentyl-1,3-dimethylxanthine (0.25 μM), a selective A₁-receptor antagonist, increased significantly the electrical field stimulation-induced release of [3H]-acetylcholine in slices prepared from 4- and 12-month-old rats, showing a tonic inhibitory action of endogenous adenosine via stimulation of presynaptic A₁-adenosine receptors. In contrast, 8-cyclopentyl-1,3-dimethylxanthine had no effect in 24-month-old rats. 2-Chloroadenosine (10 μM), an adenosine receptor agonist decreased the release of [³H]-acetylcholine in slices taken from 4- and 12-month-old rats, and no significant change was observed in slices taken from 24-month-old rats. In order to show whether the number/or affinity of the A₁-receptors was affected in aged rats, [³H]-8-cyclopentyl-1,3-dimethylxanthine binding was studied in hippocampal membranes prepared from rats of different ages. Whereas the B_max value was significantly lower in 2-year-old rats than in younger counterparts, the dissociation constant (K_d) was not affected by aging, indicating that the density rather than the affinity of adenosine receptors was altered. Endogenous adenosine levels present in the extracellular space were also measured in the superfusate by high performance liquid chromatography (HPLC) coupled with ultraviolet detection, and an age-related increase in the adenosine level was found.Insummary, our results indicate that during aging the level of adenosine in the extracellular fluid is increased in the hippocampus. There is a downregulation and reduced responsiveness of presynaptic adenosine A₁-receptors, and it seems likely that these changes are due to the enhanced adenosine level in the extracellular space.     

Postnatal changes of nicotinic acetylcholine receptor 2, α3, α4, α7 and β2 subunits genes expression in rat brain

Postnatal changes of nicotinic acetylcholine receptor (nAChR) α2, α3, α4, α7 and β2 subunits mRNAs were investigated in rat brain using ribonuclease protection assay. Multiple developmental patterns were observed: (1) transient expression during the first few postnatal weeks; α2 in the hippocampus and brain stem, α3 in the striatum, cerebellum and cortex, α4 in the hippocampus, striatum and cerebellum, α7 in the cerebellum and β2 in the striatum. (2) Constant expression across development; α2 and α3 in the thalamus, α4 in the cortex, thalamus and brain stem, α7 in the thalamus and brain stem and β2 in all brain regions except striatum. (3) Non-detection across development; α2 in the cortex, striatum and cerebellum. (4) Increase with age; α7 in the cortex and hippocampus. (5) Bell-shaped development; α7 in the striatum. Postnatal changes of nAChR isoforms in different brain regions of rat were investigated by receptor binding assays. The developmental patterns of [³H]epibatidine and (−)-[³H]nicotine binding sites were similar to each other in each brain region, but different from that of [³H]α-bungarotoxin binding sites. No obvious correlation was observed between the developmental patterns of [³H]α-bungarotoxin, [³H]epibatidine and (−)-[³H]nicotine binding sites and corresponding subunits mRNAs. These results indicate that multiple mechanisms are involved in changes of gene expression of nAChRs subunits in the brain of developing rats.     

Light-increased cGMP and K conductance in the hyperpolarizing receptor potential of Onchidium extra-ocular photoreceptors

The phototransduction mechanism of the extra-ocular photoreceptor cells Ip-2 and Ip-1 in the mollusc Onchidium ganglion was examined. Previous work showed that the depolarizing receptor potential of another extra-ocular photoreceptor cell, A-P-1 is produced by a decrease of the light-sensitive K⁺ conductance activated by a second messenger, cGMP and is inactivated by the hydrolysis of cGMP. Here, a hyperpolarizing receptor potential of Ip-2 or Ip-1 was associated with an increase in membrane conductance. When Ip-2 or Ip-1 was voltage-clamped near the resting membrane potential, light induced an outward photocurrent corresponding to the above hyperpolarization. The spectral sensitivity had a peak at 510 nm. The shift of reversal potentials of the photocurrent depended on the Nernst equation of K⁺-selective conductance. The photocurrent was blocked by 4-AP and l-DIL, which are effective blockers of the A-P-1 light-sensitive K⁺ conductance. These results suggested that the hyperpolarization is mediated by increasing a similar light-sensitive K⁺ conductance to that of A-P-1. The injection of cGMP or Ca²⁺ into a cell produced a K⁺ current that mimicked the photocurrent. 4-AP and l-DIL both abolished the cGMP-activated K⁺ current, while TEA suppressed only the Ca²⁺-activated K⁺ current. These results indicated that cGMP is also a second messenger that regulates the light-sensitive K⁺ conductance. The photocurrent was blocked by LY-83583, a guanylate cyclase (GC) inhibitor, but was unaltered by zaprinast, a phosphodiesterase (PDE) inhibitor. Together, the present results suggest that increasing the internal cGMP in Ip-2 or Ip-1 cells light-activates GC rather than inhibits PDE, thereby leading to an increase of the light-sensitive K⁺ conductance and the hyperpolarization.