Influence of sarmesin on some dopamine-related types of behaviour

Summary The present study was undertaken to investigate the effects of sarmesin, an analogue of [Sar¹] angiotensin II (ANG II) where the tyrosine hydroxyl group in position 4 is methylated, on dopamine (DA)-related paradigms: locomotor and exploratory behaviour as well as apomorphine (3 mg/kg, ip)-induced stereotypy in rats. Sarmesin (0.5 and 1 g, icv) significantly decreased ambulation and rearing movements, and blocked the inhibitory effect of ANG II (0.1 g) on both types of activity. Sarmesin induced biphasic effects on apomorphine-induced stereotypy depending on the dose increase (0.5 and 5 g, icv) and decrease (10 g). Moreover, sarmesin (5 g) blocked the inhibitory effect of ANG II (2 g, icv) on apomorphine stereotypy. Taken together, these results suggest that sarmesin might interact with AT₁ and AT₂ receptor subtypes. The results further confirm the statement for ANG II-DA interaction in brain structures involved in these types of behaviour.     

Effects of prolonged oral administration of fumonisin B1 and aflatoxin B1 in rats

The effects of prolonged oral administration (21 days) of fumonisin B₁ (FB₁) and aflatoxin B₁ (AFB₁) were evaluated on male Wistar rats. The animals were housed in individual metabolic cages and submitted to the following treatments: 1-0 g AFB₁ + 0 mg FB₁/100g bw.; 2-72 g AFB₁+ 0 mg FB₁/100 g bw; 3-0 g AFB₁ + 0.5 mg FB₁ g bw; 4-0 g AFB₁ + 1.5 mg FB₁/100 g bw; 5-72 g AFB₁ + 0.5 mg FB₁/100g bw; 6-72 gAFB₁ + 1.5 mg FB₁/100g bw. On day 21, the rats were sacrificed for evaluation. The results showed that treated animals presented differences in body weight and absolute/relative weights of liver and kidney as well as altered hepatic function and cholesterol blood levels. Rats fed with the greatest doses of AFB₁ and FB₁ gained less weight (2.79 g/day) at the end of the experimental period; their blood concentrations of liver enzymes aspartate aminotransferase (AST) and alkaline phosphatase (AP) were above control levels (130.35 /l and 471.00 /l, respectively). Blood cholesterol increased in the groups treated with the highest dose ofFB₁ or FB₁ associated with AFB₁. Histopathology revealed the occurrence of apoptosis in the liver of rats exposed to FB₁. The association of aflatoxin B₁ with fumonisin B₁ at higher dose probably potentiated the effects of the higher dose of fumonisin B₁acting singly.This revised version was published online in October 2005 with corrections to the Cover Date.     

In vitro comparison of ceftazidime,aztreonam, cefpirome,gentamicin, imipenem,enoxacin, and ticarcillin plus clavulanic acid against 108 strains ofPseudomonas maltophilia

Pseudomonas maltophilia is an uncommon cause of hospital-acquired infection and is resistant to most of the antimicrobial agents used in the treatment of gram-negative infections. Susceptibility of 108 isolates ofP. maltophilia to ceftazidime, aztreonam, defpirome, gentamicin, imipenem, enoxacin, and ticarcillin plus clavulanic acid was determined by an agar dilution method. The isolates were in general resistant to the antibiotics. Imipenem and cefpirome were not active at clinically achievable levels. Of the isolates, 20% were susceptible to 16 g/ml ceftazidime, 53% were susceptible to 4 g/ml enoxacin, 10% were susceptible to 4 g/ml gentamicin, and 25% were susceptible to 64 g/ml ticarcillin plus 2 g/ml clavulanic acid.     

Electron microscopic demonstration of phospholipase B activity in the liver and the kidney of the mouse

Summary A method has been developed for electron microscopic histochemical demonstration of phospholipase B, (lecithinase B, E C 3.1.1.5, lysolecithin acyl hydrolase), which hydrolyzes - and -positions of phospholipids in mouse liver, kidney and adrenal tissues. Tissues either fixed in cold 1% paraformaldehyde or unfixed were cut into 40 m frozen sections and were incubated at 37° C in a medium at pH 6.6 or 4.5 containing 2 M lysolecithin and 0.25 mM CaCl₂ for 20 min. The fatty acids liberated by enzymatic hydrolysis were trapped as calcium precipitate and were converted to lead precipitate by treatment with lead nitrate. The reaction products were observed by electron microscopy to be localized on the end of the smooth endoplasmic reticulum at pH 6.6 and in lysosomes and lipid droplets at pH 4.5. It is concluded that the products showed the localization of phospholipase B activity.     

Concentrations of fumonisin B1 in feeds associated with animal health problems

Ninety-eight samples of feeds associated with 44 cases of equine leukoencephalomalacia (ELEM) and 83 samples of feed associated with 42 cases of a porcine pulmonary edema syndrome (PPE) were analyzed for fumonisin B₁ (FB₁). For comparison purposes, 51 feed samples not associated with PPE or ELEM were also analyzed. Feed associated with ELEM contained FB₁ ranging from less than 1 g/g to 126 g/g with 75% of the cases having at least 1 sample above 10 g/g. Feeds associated with PPE ranged from less than 1 g/g to 330 g/g with 71% of the cases having at least 1 sample greater than 10 g/g. Quantitation was by high performance liquid chromatography (HPLC)/fluorescence using the fluorescamine derivative with confirmation by thin layer chromatography (TLC) and/or gas chromatography/mass spectroscopy (GC/MS).     

Decreased ribulose-1,5-bisphosphate carboxylase-oxygenase in transgenic tobacco transformed with “antisense” rbcS

Tobacco (Nicotiana tabacum L.) plants transformed with antisense rbcS to decrease the expression of ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) have been used to investigate the contribution of Rubisco to the control of photosynthesis in plants growing at different irradiances. Tobacco plants were grown in controlled-climate chambers under ambient CO₂ at 20°C at 100, 300 and 750 mol·m^–2·s^–1 irradiance, and at 28°C at 100, 300 and 1000 mol·m^–2·s^–1 irradiance. (i) Measurement of photosynthesis under ambient conditions showed that the flux control coefficient of Rubisco (C _infRubisco ^supA ) was very low (0.01–0.03) at low growth irradiance, and still fairly low (0.24–0.27) at higher irradiance. (ii) Short-term changes in the irradiance used to measure photosynthesis showed that C _infRubisco ^supA increases as incident irradiance rises, (iii) When low-light (100 mol·m^–2·s^–1)-grown plants are exposed to high (750–1000 mol·m^–2·s^–1) irradiance, Rubisco is almost totally limiting for photosynthesis in wild types. However, when high-light-grown leaves (750–1000 mol·m^–2·s^–1) are suddenly exposed to high and saturating irradiance (1500–2000 mol·m^–2·s^–1), C _infRubisco ^supA remained relatively low (0.23–0.33), showing that in saturating light Rubisco only exerts partial control over the light-saturated rate of photosynthesis in sun leaves; apparently additional factors are co-limiting photosynthetic performance, (iv) Growth of plants at high irradiance led to a small decrease in the percentage of total protein found in the insoluble (thylakoid fraction), and a decrease of chlorophyll, relative to protein or structural leaf dry weight. As a consequence of this change, high-irradiance-grown leaves illuminated at growth irradiance avoided an inbalance between the light reactions and Rubisco; this was shown by the low value of C _infRubisco ^supA (see above) and by measurements showing that non-photochemical quenching was low, photochemical quenching high, and NADP-malate dehydrogenase activation was low at the growth irradiance. In contrast, when a leaf adapted to low irradiance was illuminated at a higher irradiance, Rubisco exerted more control, non-photochemical quenching was higher, photochemical quenching was lower, and NADP-malate dehydrogenase activation was higher than in a leaf which had grown at that irradiance. We conclude that changes in leaf composition allow the leaf to avoid a one-sided limitation by Rubisco and, hence, overexcitation and overreduction of the thylakoids in high-irradiance growth conditions, (v) Antisense plants with less Rubisco contained a higher content of insoluble (thylakoid) protein and chlorophyll, compared to total protein or structural leaf dry weight. They also showed a higher rate of photosynthesis than the wild type, when measured at an irradiance below that at which the plant had grown. We propose that N-allocation in low light is not optimal in tobacco and that genetic manipulation to decrease Rubisco may, in some circumstances, increase photosynthetic performance in low light.Abbreviations A rate of photosynthesis - C _infRubisco ^supA flux control coefficient of Rubisco for photosynthesis - c_i internal CO₂ concentration - qE energy-dependent quenching of chlorophyll fluorescense - qQ photochemical quenching of chlorophyll fluorescence - NADP-MDH NADP-dependent malate dehydrogenase - Rubisco ribulose-1,5-bisphosphate carboxylase-oxygenase - RuBP ribulose-1,5-bisphosphate This work was supported by the Deutsche Forschungsgemeinschaft (SFB 137).     

Regeneration of plantlets through organogenesis in the Himalayan yellow poppy,Meconopsis paniculata

Plantlet formation through organogenesis in callus cultures of Himalayan yellow poppy,Meconopsis paniculata D.Don (Prain), a threatened taxon of ornamental value, is described. Hypocotyl segments from 3-month-old laboratory-raised seedlings produced callus on agar-solidified Murashige and Skoog medium (MS) containing 10 M -naphthaleneacetic acid and 1 M kinetin. Shoots differentiated best from callus on MS containing 10 M indolebutyric acid (IBA) and 1 M 6-benzyladenine. The regenerated shoots rooted best on MS medium containing 10 M IBA. From seed germination to differentiation of plantlets through the two-step organogenesis process required 28–29 weeks.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - FAA formalin-acetic acid-alcohol - BA 6-benzyladenine - IAA indole-acetic acid - IBA indolebutyric acid - GA₃ gibberellic acid - NAA -naphthaleneacetic acid - RH relative humidity     

Effects of GABA antagonists,SR 95531 and bicuculline,on GABAA receptor-regulated chloride flux in rat cortical synaptoneurosomes

Synaptoneurosomes isolated from cerebral cortices of male Sprague-Dawley rats were used for studying GABA_A receptor-regulated chloride influx. The in vitro effects of GABA antagonists, SR 95531 (a pyridazinyl GABA derivative) and bicuculline, on pentobarbital-stimulated, muscimol-stimulated or flunitrazepam-enhanced, muscimol-stimulated chloride uptake were studied. The chloride uptake was determined at 30°C, for 5 sec. Pentobarbital and muscimol produced a maximal stimulation of chloride uptake in cortical synaptoneurosomes at 500 M and 50M, respectively. SR 95531 as well as bicuculline had no effect on the basal uptake of chloride. Whereas, SR 95531 (0.3–30 M) and bicuculline (0.1–100 M), when added 5 min before muscimol (50 M), produced a significant concentration-dependent inhibition of muscimol (50 M)-stimulated chloride uptake (IC₅₀ s of 0.89±0.11 M and 13.45±2.10M, respectively). In studies of the inhibitory effects of SR 95531 and bicuculline on pentobarbital (500 M)-stimulated chloride uptake, the IC₅₀ s were 0.81±0.12 M and 3.86±1.14 M, respectively. SR 95531 exhibited a more potent inhibitory effect than bicuculline on flunitrazepam-enhanced, muscimol-stimulated chloride uptake. The results revealed that SR 95531 has a more potent antagonistic effect than bicuculline on GABA_A-regulated chloride flux.     

Aflatoxin influences on seed germination and root elongation by two cultivars of Glycine max and uptake of 65 Zn-ZnCl2 by the cultivars

Maximum inhibition of Glycine max, cv. Essex seed germination occurred at 10 g/ml following 72 hr imbibition in constant light. Seeds imbided 108 hr in constant darkness at this concentration showed a 20% rise in germination over that of the control. Imbibition of G. max, cv. Williams seeds in either light or dark for 96 hr did not suppress germination. Imbibition of Essex seeds in either light or dark at 2.5 through 10 g/ml stimulated root elongation except for 10 g/ml at 96 hr (light). Maximum inhibition of Williams root elongation under constant light was at 48 and 72 hr with 10 g/ml. Statistically significant differences in cotyledon, leaf and stem lengths between non-treated (NT) and treated (T) seedlings were not found except for Williams stem length at 2.5 / ml. Root elongation was stimulated 1.2- and 1.1-folds, respectively, at 5.0 (Essex) and 2.5 (Williams) g/ml. Toxin at 2.5 through 10.0 g/ml did not markedly alter either cotyledon or leaf widths with the exception of Williams leaf width at 2.5 g/ml. Medium supplementation with 2.5 through 10.0 g/ml resulted in cotyledon, leaf and root weight enhancements for Essex seedlings. Stem weight was not markedly affected. An 18% rise in Williams cotyledon weight above that of the control was seen at 2.5 g/ml. Williams leaf weights were increased 1.75- and 1.25-folds, respectively, at 2.5 and 10.0 g/ml. Aflatoxin B₁, at 2.5 g/ml promoted Williams stem and root elongation 1.20- and 1.09-folds, respectively. Most of the radioactivity from ⁶⁵Zn-ZnCl₂ recovered within organs was found within Essex roots for both T and NT seedlings. A higher amount of radioactivity was recovered within roots at each toxin concentration than was without toxin. However, this was not statistically significant. Significant differences in the distribution of radioactivity within roots between NT and T Williams seedlings were not observed. Generally, AFB₁ failed to affect significantly these two varieties of soybeans based on the tests relating to germination, growth and radiolabel uptake.     

Effects of polymyxin B on the sarcoplasmic reticulum membrane

Polymyxin B, a cyclic peptide antibiotic, inhibits Ca²⁺-ATPase, p-nitrophenyl phosphatase and phosphorylase kinase activities associated with rabbit skeletal muscle sarcoplasmic reticulum membranes; 50% inhibition is induced by 100 M, 130M and 550 M of polymyxin respectively. The fluorescence intensity of fluorescein isothiocyanate-labeled Ca²⁺-ATPase, decreases in the presence of polymyxin (50% of the total decrease at 70 M polymyxin). On the other hand, the polypeptide inhibits calmodulin-dependent endogenous phosphorylation of 60 kDa, 20 kDa and 14 kDa membrane proteins, while an increase of calmodulin-dependent phosphorylation is observed in 132 kDa and 86 kDa proteins.     

Direct somatic embryogenesis through thin cell layer culture in Panax ginseng

Direct somatic embryos were differentiated on cotyledon transverse Thin Cell Layers (tTCLs) of Panax ginseng after 9 weeks in the Murashige and Skoog basal (MS) medium containing 2,4-d (5M). When MS medium containing 2,4-d (5M) was used for seedling pretreatment and for tTCLs culture, somatic embryos were observed 2 weeks earlier, i.e. after 7 weeks of culture. On the tTCLs from seedlings pretreated with 2,4-d (5M) combined with benzyladenine and zeatin at 0.1 M (BZ), somatic embryos were observed after 6 weeks of culture and the percentage of embryogenesis was higher (62%) than when 2,4-d was used alone for pretreatment (40%). Similar results were also obtained from pretreatment with combinations of 2,4-d (5M) and thidiazuron (TDZ) (0.01, 0.1M). When a combination of 2,4-d (5M) and BZ (0.1M) was used both for seedling pretreatment and for tTCLs culture, both somatic embryos and shoots were observed after only 3 weeks. As the concentration of BZ increased, the percentage of somatic embryogenesis decreased but the percentage of organogenesis increased. Similar responses were obtained with a combination of 2,4-d (5M) and TDZ (0.01M). On the medium containing both NAA (0.3M) and BZ (1M), globular- and heart- stage embryos developed after 4 weeks of culture into cotyledonary-staged embryos which remained dormant after a short elongation of the embryo axis. The importance of seedling pretreatment by growth substances in enhancing somatic embryogenesis is reported.Abbreviations BA 6-benzyladenine - BZ combination of BA and zeatin - 2,4-d 2,4-dichlorophenoxyacetic acid - MS medium Murashige and Skoog basal medium - NAA a-naphthaleneacetic acid - TDZ thidiazuron - tTCLs transverse thin cell layers - TCL longitudinal thin cell layer     

Fumonisin and Beauvericin Induce Apoptosis in Turkey Peripheral Blood Lymphocytes

Fumonisins, a family of mycotoxins produced by Fusarium verticillioides (synonym Fusarium moniliforme Sheldon) and F. proliferatum, have been associated with various deleterious effects in different animal species. Serological, hematological and pathological effects and mortality have previously been observed in broiler chicks fed F. proliferatum culture material containing known concentrations of fumonisin, moniliformin and beauvericin. Turkey peripheral blood lymphocytes were exposed in vitro for 72 hours to fumonisin B₁(FB₁), fumonisin B₂(FB₂), hydrolyzed fumonisin B₁ (HFB₁), moniliformin and tricarballylic acid (TCA) (0.01-25 g/ml). A decrease in cell proliferation, as determined by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] bioassay, occurred in the order: FB₂ > FB₁ > HFB₁, with IC₅₀ = 0.6 M, 1 M and 10 M, respectively. Internucleosomal DNA fragmentation and morphological features characteristic of apoptosis were observed following exposure to fumonisin B₁ and beauvericin; cytoplasmic condensation and membrane blebbing were seen by light microscopy. Tricarballylic acid and moniliformin did not interfere with cell proliferation. Results suggested that fumonisin B₁ and beauvericin may affect immune functions by suppressing proliferation and inducing apoptosis of lymphocytes.     

Tissue culture ofKarwinskia humboldtiana—a plant producing toxins with antitumoural effects

Isolated embryos ofKarwinskia humboldtiana were cultured in vitro. The growth of embryos and development to plantlets on woody plant medium supplemented with indole-3-acetic acid 6.10^-2 mol l^–1, gibberellic acid (GA₃) 3.10^-2 mol l^–1, and 6-benzylaminopurine (BA) 2 mol l^–1 was obtained. Multiplication of shoots and rooting of excised shoots has been achieved. Callus formation on modified Murashige-Skoog medium supplemented with 1-naphthaleneacetic acid 10 mol l^–1, GA₃ 14 mol l^–1, and kinetin 5 mol l^–1 on hypocotyls, or on root cultures on medium supplemented with 2.4-dichlorophenoxyacetic acid 10 mol l^–1 and BA 10 mol l^–1 was induced.Abbreviations BA 6-benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - TEM transmission electron microscopy     

Somatic embryogenesis and plant regeneration inMedicago suffruticosa

A successful procedure has been designed for the regeneration of plantlets from leaf sections of the self-pollinating species,Medicago suffruticosa. Callus growth was promoted by a 4-week culture period on liquid Kao's medium containing 4.9 M benzyladenine and 4.5 M 2,4-dichlorophenoxyacetic acid (2,4-d), followed by a 4-day treatment in which the benzyladenine was elevated to 44.4 M. Shoots/plantlets were observed after 3–4 weeks culture on growth regulator-free agar-solidified medium. Under these conditions, the regeneration frequency from callus was 18% and a histological study showed that this regeneration was through somatic embryogenesis. The growth regulator treatment, with a relatively high concentration of growth regulators (44.4 M benzyladenine) for a short time period (4 days), is important for inhibiting polyphenol compounds and for stimulating callus growth and plant regeneration.Abbreviations 2,4-d 2,4-dichiorophenoxyacetic acid - BA benzyladenine - NAA -napthaleneacetic acid     

Male reproductive effect of arsenic in mice

Arsenic, a known human carcinogen, was given to mice via drinking water as sodium arsenite at a dose 53.39, 133.47, 266.95 and 533.90 mol l for 35 days. A decrease in the activity of 17 HSD along with increase in LDH, GT activity were observed at 533.90 mol l. The observed sperm count, motility and morphological abnormalities in sperm were similar to control at lower dose levels. However at 533.90 mol l a significant decrease in sperm count and motility along with increase in abnormal sperm were noticed. Significant accumulation of arsenic in testes and accessory sex organs may be attributed to the arsenic binding to the tissues or greater cellular uptake. No effects were observed on indices studied for reproductive effects at 53.39 mol l arsenic close to which human being are exposed through drinking water under the present set of experimental conditions.     

Indole butyric acid induces regeneration of phenotypically normal apricot (Prunus armeniaca L.) plants from immature embryos

A prerequisite for most transformation systems is an efficient and reliable method to regenerate phenotypically normal plants. Immature embryos or cotyledons were cultured at three developmental stages (stage 1, 2 and 3, PF=3, 30–60, and 100, respectively) from two unrelated apricot genotypes, Zard and NJA82. Explants were cultured on MS media supplemented with either BA or TDZ at four concentrations (0, 0.5, 5.0 or 20 M) and 2,4-D at 0 or 1 M. Stage 1 embryos cultured on MS medium without growth regulators formed embryoid-like structures. Shoot primordia induction was greatest with stage 2 cotyledons on media containing 5–20 M TDZ and 1 M 2,4-D, although shoot morphology was abnormal, especially with the highest level of TDZ. In another factorial experiment, stage 2 cotyledons were cultured on media containing TDZ (0, 5, 7.5, 10, 15 or 20 M) in combination with either no auxin, 1 M 2,4-D, 1 M IBA, or 5 MIBA. Regeneration percentages of 80% or more were observed on media containing 1–5 M IBA and 5–10 M TDZ. The medium containing 5 M IBA and no TDZ exhibited the highest frequency of phenotypically normal plantlet regeneration.Abbreviations BA 6-benzylaminopurine - IBA indole-3-butyric acid - 2,4-D 2,4 dichlorophenoxyacetic acid - 2iP 2-isopentenyladenine - MS Murashige and Skoog basal medium - NAA 1-naphthaleneacetic acid - PF percent fill [(embryo length/seed length) × 100] - TDZ thidiazuron [N-phenyl-N(1,2,3,-thidiazol-5-yl)-urea] - WPM McCown's woody plant medium     

Direct Organogenesis from Mature Leaf and Petiole Explants of Eryngium Foetidum L.

Eryngium foetidum L. plants were regenerated from mature leaf and petiole explants through direct organogenesis without intervening callus phase. From leaf explants, adventitious multiple shoots raised on Murashige and Skoog (MS) medium supplemented with 4.43 M benzylaminopurine (BAP) and 0.57 M indole-3-acetic acid (IAA), whereas in petiole explants shoot regeneration occurred at 8.86 M BAP and 0.57 M IAAA. 80% of the leaf explants and 44% of petiole explants produced shoots after four weeks of culture. The regenerated plants were rooted on MS medium supplemented with 2.46 M indole-3-butyric acid and 2.88 M gibberellic acid. The plants were successfully established in the soil and showed 70.9% survival in the field.     

Clozapine and Several Other Antipsychotic/Antidepressant Drugs Preferentially Block the Same ‘Core’ Fraction of GABAA Receptors

Clozapine and several other antipsychotic/antidepressant drugs that fully or partially block GABA_A receptors were tested at concentrations that reversed the inhibitory effect of 1 M GABA on ³⁵S-t-butylbicyclophosphorothionate ([³⁵S]TBPS) binding to rat forebrain membranes only about 20–30%, here designated core fractions. Clozapine at 10 M reverses 1 M GABA 25 ± 4.0% (n = 23) (its core fraction). Fourty three compounds were tested alone, and pairwise together with 10 M Clozapine. The core fractions of some of the compounds yielded significant additive reversals together with 10 M Clozapine, while others did not. A group of 14 compounds of which 7 are clinically effective antipsychotic drugs, including Chlorprothixene, Clomacran, Clopipazan, Fluotracen, Sulforidazine, Thioproperazine, and cis-Thiothixene, were statistically non-additive with 10 M Clozapine, suggesting that all of these drugs selectively block the same core population of GABA_A receptors as Clozapine. These non-additivities also suggest that Clozapine at 10 M fully saturates a subset of GABA_A receptors blocked by 1 M GABA. Therefore, Clozapine probably blocks 2 or more types of GABA_A receptors, but only half of the receptors that are sensitive to 1 M GABA. A second group of 12 compounds of which 6 are clinically active antidepressant/antipsychotic drugs including Amoxapine, Clothiapine, Dibenzepine, Inkasan (Metralindole), Metiapine and Zimelidine were slightly, but significantly, additive with Clozapine suggesting that these compounds block most of Clozapine's core fraction, plus a small additional fraction. A third group consisted of ten compounds that yielded larger (R > 80) and statistically highly significant additivities with Clozapine. Complete additivity was obtained with Bathophenanthroline disulfonate, and Isocarboxazid, suggesting that they block GABA_A receptors other than those blocked by 10 M Clozapine. Seven classical GABA_A receptor blockers, also tested at concentrations yielding 21 to 33% reversal alone, were all significantly additive with 10 M Clozapine, but in no case was the additivity complete. The largest additivity was obtained with Pitrazepine (21%) and the smallest with Tubocurarine (9%). These results provide further support for the notion that selective blockade of the same subset of GABA_A receptors may contribute to the clinical antipsychotic/antidepressant effects of Clozapine. The B_opt values for Clozapine are 50 ± 1.7% and 26 ± 2.6% ( n = 3) in whole rat forebrain and cerebellum, respectively, confirming that clozapine-sensitive GABA_A receptors are unevenly distributed in the brain. The sedative and anxiolytic properties of Clozapine and other antipsychotic drugs may be due to selective blockade of GABergic disinhibition at certain interneurons.     

Regeneration of pea (Pisum sativum L. cv. Century) plants by in vitro culture of immature leaflets

In vitro regeneration of plants from immature leaflets of 3 day-old pea (Pisum sativum L. cv. Century) seedlings was studied under defined nutritional, hormonal and environmental conditions. Immature leaflets isolated from the second and third apical leaves of aseptically germinated seeds were cultured on MS medium containing vitamins as in B5 medium, 3% sucrose, 0.8% agar and supplemented with 0.1, 1, and 10 M concentrations of naphthaleneacetic acid (NAA) and 1 and 10 M levels of benzyladenine (BA) in various combinations. Shoot regeneration from the primary callus occurred within 45 to 90 days of culture in most of the hormone combinations. Although the number of calli producing shoots was maximal at 10 M levels of NAA and BA, multiple shoot regeneration was predominant at a combination of 0.1 M NAA and 10 M BA. Indoleacetic acid (IAA) and kinetin (K), both at 10 M, also induced shoot regeneration. No shoots were regenerated when 10 day-old leaflets were used as explants. Root production generally occurred on non-shoot regenerating calli. Roots were induced to differentiate by transferring the regenerated shoots onto half-strength B5 medium supplemented with 1 M NAA.NRCC # 19712Visiting Scientist, supported by a research fellowship from Consejo Nacional de Investigaciones Científicas y Técnicas (Rep. Argentina). Permanent address: Facultad de Ciencias Agrarias, IBONE, Casilla de Correos 209, Corrientes (3400), Argentina.     

Rapid plant regeneration of pea using thidiazuron

A simple and rapid pea regeneration procedure was developed. An average of up to 20 shoots formed from hypocotyl explants of cvs. Sugar Ann and Patriot cultured on Murashige and Skoog basal medium supplemented with 0.5 or 1.0 M thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea). Hypocotyls of Puget and Sugar Daddy did not respond. Regenerated shoots rooted rapidly when cultured on Murashige and Skoog basal medium containing either 2.0 M -naphthaleneacetic acid or 1.0–2.0 M indole-3-butyric acid. Seeds were harvested from regenerated plants after only 9–11 weeks.Abbreviations BAP 6-benzyladenine - 2,4-d 2,4-dichlorophenoxy acetic acid - GA₃ gibberellic acid - IBA indole-3-butyric acid - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic acid - TDZ thidiazuron (N-phenyl-N-1,2,3-thiadiazol-5-ylurea)