Evidence that 5-HT2 antagonism elicits a 5-HT3-mediated increase in dopamine transmission

Amperozide, a novel atypical antipsychotic drug with few extrapyramidal side effects, is a strong serotonin₂ (5-HT₂) antagonist but has low affinity for dopamine receptors in vitro. The effect of amperozide on the dopaminergic synapse was studied with an in vivo microdialysis technique using anesthetized male Sprague-Dawley rats. Following implantation of dialysis probes into the striatum and nucleus accumbens (NuAc), amperozide was intravenously infused as six consecutive incremental doses (0.5, 0.5, 1.0, 2.0, 4.0 and 8.0 mg/kg) at intervals of 15 min. From the beginning of drug infusion, perfusates were collected in fractions every 30 min throughout a total period of 120 min. The samples were then immediately analyzed by high-performance liquid chromatography with electrochemical detection. Amperozide induced a dose-related elevation of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindolacetic acid (5-HIAA) levels in both areas.p-Chlorophenylalanine (pCPA) pretreatment abolished the production of 5-HIAA in both areas and attenuated the amperozide-induced rise of DOPAC but not of dopamine. After pretreatment with an intravenous 5-HT₃ antagonist, MDL 72222, the amperozide-induced changes in dopamine, DOPAC and 5-HIAA in both areas were lower than in the saline control group. Preliminary data showed that afterpCPA pretreatment, incremental concentrations of the 5-HT₃ agonist 1-(m-chlorophenyl)-biguanide perfused via the probe also produced significant elevation of dopamine and DOPAC levels in these two areas. Taken together, these results suggest that amperozide may directly block 5-HT₂ receptors in the striatum and NuAc, thereby enhancing 5-HT transmission. The enhanced 5-HT transmission may activate postsynpatic 5-HT₃ receptors located on the dopaminergic terminals, leading to changes in dopamine transmission in these two areas.     

Localization of proteasomes in plant cells

Summary Proteasomes, also known as multicatalytic proteinase complexes, were localized in suspension cells of potato (Solanum tuberosum) by direct immunofluorescence using polyclonal antibodies labelled with fluorescein isothiocyanate. The method used allows an estimate of relative amounts of proteasomal antigens in different cell components. Proteasomes are present in the nuclei and the cytoplasm. The nucleoplasm contains small areas of weak fluorescence. The peripheral cytoplasm and possibly elements of the cytoskeleton show higher fluorescence than other parts of the cytoplasm. This indicates a localization of proteasomes similar to that known from animal cells.Abbreviations DMSO dimethylsulfoxide - EGTA ethyleneglycol-bis-(-aminoethylether)-N,N,N,N-tetra acetic acid - FITC fluorescein isothiocyanate - PBS phosphate buffered saline - PIPES piperazine-1,4-bis-(2-ethanesulfonic acid)     

Modulation by angiotensin III of nociception-related and arterial pressure-related neuronal responsiveness in the nucleus reticularis gigantocellularis of the rat

We evaluated possible modulation by angiotensin III (AIII) of the interactive effect of noxious stimuli and elevation in systemic arterial pressure on the responsiveness of neurons in the nucleus reticularis gigantocellularis (NRGC) of the medulla oblongata. Combined extracellular single-neuron recording and microiontophoresis were carried out on male, adult Sprague-Dawley rats anesthetized with pentobarbital sodium. The responsiveness of NRGC neurons to nociception (tail clamp) and/or transient hypertension elicited by phenylephrine (5 μg/kg, i.v.), in the absence or presence of AIII, was used as the experimental index. Microiontophoretic application of the heptapeptide suppressed the responses of spontaneously active NRGC neurons to individually delivered nociception or hypertension. Interestingly, the preferential reduction in responsiveness to tail clamp upon simultaneous elevation in arterial pressure was reversed to one that favored nociception in the presence of AIII. These actions of the heptapeptide appeared to be receptor-specific, since they were discernibly blocked by its selective antagonist, Ile⁷-angiotensin III. Our results reveal that neuropeptides such as AIII may differentially modulate neuronal responsiveness according to the prevailing physiologic input(s) to the central nervous system of the animal.     

Stress-Induced Sensitization of Dopamine and Norepinephrine Efflux in Medial Prefrontal Cortex of the Rat

Abstract: We examined whether prior exposure to chronic cold (17–28 days, 5°C) alters basal or stress-evoked (30-min tail shock) catecholamine release in medial prefrontal cortex, nucleus accumbens, and striatum, using in vivo microdialysis. Basal norepinephrine (NE) concentrations in medial prefrontal cortex did not differ between chronically cold-exposed rats and naive control rats (2.7 ± 0.3 vs. 2.5 ± 0.2 pg/20 µl, respectively). Basal dopamine (DA) efflux in any of the brain regions was not significantly different between chronically cold-exposed rats and naive rats. However, a trend for lower basal DA efflux in the cold-exposed relative to naive rats was observed in medial prefrontal cortex (1.5 ± 0.2 vs. 2.2 ± 0.3 pg/20 µl, respectively), nucleus accumbens (3.7 ± 0.8 vs. 5.4 ± 0.9 pg/20 µl, respectively), and striatum (4.4 ± 0.5 vs. 7.2 ± 1.5 pg/20 µl, respectively). In medial prefrontal cortex of rats previously exposed to cold, tail shock elicited a greater increase from baseline in both DA and NE efflux relative to that measured in naive rats (DA, 2.3 ± 0.3 vs. 1.2 ± 0.1 pg, respectively; NE, 3.8 ± 0.4 vs. 1.4 ± 0.2 pg, respectively). However, in nucleus accumbens or striatum of rats previously exposed to cold, the stress-induced increase in DA efflux was not significantly different from that of naive rats (nucleus accumbens, 1.8 ± 0.7 vs. 1.5 ± 0.3 pg, respectively; striatum, 1.9 ± 0.4 vs. 2.6 ± 0.7 pg, respectively). Thus, both cortical NE projections and cortically projecting DA neurons sensitize after chronic exposure to cold. In contrast, subcortical DA projections do not sensitize under these conditions.     

Clearance of Exogenous Dopamine in Rat Dorsal Striatum and Nucleus Accumbens: Role of Metabolism and Effects of Locally Applied Uptake Inhibitors

In vivo electrochemistry was used to investigate the mechanisms contributing to the clearance of locally applied dopamine in the dorsal striatum and nucleus accumbens of urethane-anesthetized rats. Chronoamperometric recordings were continuously made at 5 Hz using Nafion-coated carbon fiber electrodes. When a finite amount of dopamine was pressure-ejected at 5-min intervals from a micropipette adjacent to the electrode, transient and reproducible dopamine signals were detected. Substitution of L-a-methyldopamine, a substrate for the dopamine transporter but not for monoamine oxidase, for dopamine in the micropipette did not substantially alter the time course of the resulting signals. This indicates that metabolism of locally applied dopamine to 3,4-dihydroxyphenylacetic acid is not responsible for the decline in the dopamine signal. Similarly, changing the applied oxidation potential from ±0.45 to ±0.80 V, which allows for detection of 3-methoxytyramine formed from dopamine via catechol-O-methyltransferase, had little effect on signal amplitude or time course. In contrast, lesioning the dopamine terminals with 6-hydroxydopamine, or locally applying the dopamine uptake inhibitors cocaine or nomifensine before pressure ejection of dopamine, significantly increased the amplitude and time course of the dopamine signals in both regions. The effects of cocaine and nomifensine were greater in the nucleus accumbens than in the dorsal striatum. Local application of lidocaine and procaine had no effect on the dopamine signals. Initial attempts at modeling resulted in curves that were in qualitative agreement with our experimental findings. Taken together, these data indicate that (1) uptake of dopamine by the neuronal dopamine transporter, rather than metabolism or diffusion, is the major mechanism for clearing locally applied dopamine from the extracellular milieu of the dorsal striatum and nucleus accumbens, and (2) the nucleus accumbens is more sensitive to the effects of inhibitors of dopamine uptake than is the dorsal striatum.     

Cell division in caffeine-induced binucleate protonemal cells ofAdiantum. II. Formation of the preprophase band and cell division in centrifuged and non-centrifuged cells

Organization of microtubules (MTs) in relation to the behavior of nuclei was examined in dividing binucleate cells ofAdiantum capillus-veneris L. To induce binucleate cells, caffeine, an inhibitor of formation of the cell plate, was applied at 4 mM to synchronously dividing protonemal cells during cytokinesis (Murata and Wada 1993). Formation of the preprophase band (PPB) during the next cell cycle was examined in non-centrifuged and centrifuged cells. The two nuclei were separated or associated with one another in both non-centrifuged and centrifuged cells, although the location of the nuclei in the cylindrical protonemal cells was different (Murata and Wada 1993). Irrespective of centrifugation, a single PPB was formed around the nuclei in cells with associated nuclei. Two PPBs were formed in cells with separated nuclei in centrifuged cells. Patterns of mitosis and cytokinesis varied, depending on the location of the PPB and the distribution of the nuclei. The role of the nucleus in formation of the PPB is discussed.     

Further characterization of nociception-related and arterial pressure-related neuronal responses in the nucleus reticularis gigantocellularis of the rat

The present study was undertaken to further characterize the nucleus reticularis gigantocellularis (NRGC) of the medulla oblongata in the central processing of nociceptive and cardiovascular signals, and its modulation by metenkephalin. In Sprague-Dawley rats anesthetized with pentobarbital sodium, we found that all 125 spontaneously active NRGC neurons that responded to noxious stimuli (tail clamp) also exhibited arterial pressure-relatedness. Forty neurons additionally manifested cardiac periodicity that persisted even during nociceptive responses. While maintaining their cardiovascular responsive characteristics, the nociception-related NRGC neuronal activity was blocked, naloxone-reversibly (0.5 mg/kg, i.v.), by morphine (5 mg/kg, i.v.). Microiontophoretically applied met-enkephalin suppressed the responsiveness of NRGC neurons to individually delivered tail clamp or transient hypertension induced by phenylephrine (5 µg/kg, i.v.). Interestingly, in NRGC neurons that manifested both nociception and arterial pressure relatedness, the preferential reduction in the response to noxious stimuli upon simultaneous elevation in systemic arterial pressure was reversed to one that favored nociception in the presence of met-enkephalin. All actions of met-enkephalin were discernibly blocked by the opioid receptor antagonist, naloxone. Our results suggest that individual NRGC neurons may participate in the processing of both nociceptive and cardiovascular information, or in the coordination of the necessary circulatory supports during nociception. In addition, neuropeptides such as met-enkephalin may exert differential modulation on neuronal responsiveness according to the prevailing physiologic status of the animal. They also showed that NRGC may be a central integrator for pain and cardiovascular-related functions.     

First divisions of somatic embryogenic cells inCichorium hybrid “474”

Summary First divisions of embryogenic cells were studied in leaves of plantlets of aCichorium hybrid (C. intybus L. ×C. endivia L.) cultured in vitro in a liquid agitated medium, at 35 °C in the dark. Stages of reactivation of competent cells were characterized by increase of nucleus and nucleolus diameter, migration of the nucleus in the centre of the cell and thickening of the cell wall. The first division of reactivated embryogenic cells was symmetrical and anticlinal in regard to the xylem vessels orientation. Embryogenic structures consisted in I-type tetrads or in rows of 4–8 cells. Then the divisions occurred in thickness at one end, without polarization or formation of a suspensor-like structure.Abbreviations EC embryogenic cell - ES embryogenic structure     

Pervasive migration of organellar DNA to the nucleus in plants

A surprisingly large number of plant nuclear DNA sequences inferred to be remnants of chloroplast and mitochondrial DNA migration events were detected through computer-assisted database searches. Nineteen independent organellar DNA insertions, with a median size of 117 by (range of 38 to >785 bp), occur in the proximity of 15 nuclear genes. One fragment appears to have been passed through a RNA intermediate, based on the presence of an edited version of the mitochondrial gene in the nucleus. Tandemly arranged fragments from disparate regions of organellar genomes and from different organellar genomes indicate that the fragments joined together from an intracellular pool of RNA and/or DNA before they integrated into the nuclear genome. Comparisons of integrated sequences to genes lacking the insertions, as well as the occurrence of coligated fragments, support a model of random integration by end joining. All transferred sequences were found in noncoding regions, but the positioning of organellar-derived DNA in introns, as well as regions 5 and 3 to nuclear genes, suggests that the random integration of organellar DNA has the potential to influence gene expression patterns. A semiquantitative estimate was performed on the amount of organellar DNA being transferred and assimilated into the nucleus. Based on this database survey, we estimate that 3–7% of the plant nuclear genomic sequence files contain organellar-derived DNA. The timing and the magnitude of genetic flux to the nuclear genome suggest that random integration is a substantial and ongoing process for creating sequence variation.Correspondence to: J.L. Blanchard