Effect of temporal synergism of neural oscillations on photorefractoriness in Japanese quail (Coturnix coturnix japonica)

Circadian rhythms in many metabolic functions including neural (transmitters) and hormonal secretion appear to change with physiological condition. It is also reported that seasonal changes in photoperiodism/reproduction and other metabolic conditions may result from a temporal interaction of circadian neural oscillations that change seasonally. To test this hypothesis, the present study was designed to study the effect of temporal synergism of two neural oscillations (serotonin and dopamine) on relative photorefractoriness of Japanese quail.Serotonin and dopamine precursor drugs (5-HTP, 5-hydroxytryptophan and L-DOPA, L-dihydroxyphenylalanine) were administered (intraperitonially 5 mg/100 g body weight) at six different time intervals of 0, 4, 8, 12, 16 and 20 hr in sexually mature quail (>12 weeks old). The birds of control group received two daily injections of normal saline. The treatment was given for 13 days in continuous condition of light and then the quail were shifted to intermediate daylength (LD 13.5:10.5 for experiment 1) and short daylength (LD 8:16 for experiment 2). Six weeks following treatment, birds in intermediate daylength showed regressed cloacal gland and testicular activity except in 12-hr group, which exhibited gonadostimulatory condition. But birds of all the groups in short daylength showed complete regression of cloacal gland after 4 weeks of the treatment. In experiment 3, reproductively quiescent relative photorefractory quail maintained under intermediate daylength (LD 13.5:10.5) received 13 daily injections of 5-HTP and L-DOPA at the interval of 12 hr. At 6 weeks post-treatment, it was observed that unlike cloacal gland of control quail, which remained regressed, that of 12-hr quail showed significant development.These findings indicate that 12-hr temporal interaction of 5-HTP and L-DOPA administration maintained reproductive system in stimulated condition and prevented reproductive regression in photorefractory quail, but did not prevent the onset of scotosensitivity. It is concluded that the 12-hr temporal relationship of circadian serotonergic and dopaminergic oscillations not only eliminates photorefractoriness but may also re-establish photosensitivity in relative photorefractory quail. These findings suggest the regulatory role of neural oscillations and their temporal interaction in the regulation of neuroendocrine-gonadal axis with special reference to photosensitivity/refractoriness.     

Effect of 12 hr temporal relation of serotonin and dopamine precursor drugs (5-HTP and L-DOPA) on photosexual responses of immature Japanese quail

To study the interaction of photoperiod and circadian neurotransmitter activity, L-DOPA L-dihydroxyphenylalanine, dopamine precursor) was administered daily, 12 hr after 5-HTP (5-hydroxy tryptophan, serotonin precursor) in sexually immature Japanese quail, raised under short photoperiod (LD8:16) since hatching. The 12 hr treatment of 5-HTP and L-DOPA was given under continuous condition of light for 11 days. After treatment the quail were transferred to intermediate day length (LD 13.5:10.5). The cloacal gland size of drug treated group increased significantly in comparison to control. The quail were then transferred to short photoperiod (LD 8:16). The cloacal gland size of both the groups started decreasing gradually but the rate was significantly low in drug treated quail in comparison to control. The results indicate that the endogenous mechanism controlling seasonality may be reset by drugs that influence serotonergic and dopaminergic activity. The 12 hr relation between the two drugs is stimulatory for gonadal growth under intermediate day length and retards the rate of regression when transferred to short days.     

Temporal synergism of neurotransmitters (serotonin and dopamine) affects testicular development in mice

The temporal phase relation of circadian oscillations is reported to regulate reproduction in many seasonally breeding avian and mammalian species, but its role in the reproductive regulation of continuous breeders is not yet known. Hence in the present study, six experimental groups of 3-week-old male Parkes strain mice, Mus musculus, were injected with 5-hydroxytryptophan (5-HTP, serotonin precursor) and l-dihydroxyphenylalanine (l-DOPA, dopamine precursor) at intervals of 0, 4, 8, 12, 16 or 20 hr (5 mg/100 g body weight per day for 13 days). Control mice received two daily injections of normal saline. When observed 24 days post-treatment, 8-hr mice exhibited low body weight and suppression of gonadal activity (spermatogenesis, sperm count/motility/viability and plasma testosterone concentration), while body weight and degree of gonadal development were higher in the 12-hr mice as compared to the controls. It is concluded that normal somatic and gonadal growth of pre-puberal mice may be suppressed with an 8-hr phase relation of circadian serotonergic and dopaminergic oscillations. On the other hand, a 12-hr phase relation accelerated the rate of gonadal maturation, while other relations led to more or less similar gonadal development as in the control mice. This study suggests the importance of circadian organization as a function of specific temporal phase relations of neural oscillations in the maturation of gonads. Although the exact mechanism still needs to be investigated, this seems to be mediated via effects on the neuroendocrine axis.     

Effects of Simulated Hypo‐ and Hyper‐Reproductive Conditions on the Characteristics of Circadian Rhythm in Hypothalamic Concentration of Serotonin and Dopamine and in Plasma Levels of Thyroxine,Triiodothyronine, and Testosterone in Japanese Quail,Coturnix coturnix japonica

In this study, hypo‐ and hyper‐reproductive conditions, as measured by concentrations of plasma testosterone in male Japanese quail held on long days LD 16:8, were experimentally simulated with injections of 5‐hydroxytryptophan (5‐HTP) and L‐dihydroxyphenylalanine, (L‐DOPA) with 8 h and 12 h phase angle differences between them in intact and melatonin‐treated birds. The effects of these treatments were assessed on the characteristics of the circadian rhythm in the hypothalamic concentration of serotonin (5‐HT), dopamine (DA), and plasma levels of thyroxine (T₄), triiodothyronine (T₃), and testosterone (T). These rhythms were also studied in sham‐operated (SO), pinealectomized (Px), vehicle‐ (Veh), and melatonin (Mel)‐treated birds. On the basis of the circadian mesors of the testosterone rhythms, three distinct categories could be identified: category A (i.e., normal breeding concentrations of plasma testosterone), which includes control, sham‐operated, and vehicle‐treated groups; category A⁺ (i.e., concentrations of plasma testosterone higher than that found in normal breeding quail), which includes 12 h, 12 h+vehicle‐treated, and Px quails; and category A^? (concentrations of plasma testosterone lower than that found in normal breeding quail), which includes 8 h, melatonin‐, and 12 h+melatonin‐treated groups. It is evident that in normal and hypergonadal conditions (i.e., birds belonging to categories A and A⁺) the circadian rhythm in hypothalamic serotonin maintained a positive phase angle of about 16 h. In contrast, birds of category A^? (i.e., in a hypogonadal condition) exhibited a negative phase angle of about 2 h. The present results clearly suggest that the internal phase relationship between the circadian rhythms in hypothalamic serotonin and dopamine might play a crucial role in strategizing and conferring a particular reproductive status to the birds. The role of circadian mechanisms involving circulating thyroid hormones in conferring reproductive status is completely ruled out, as no definite internal phase angle between these two hormonal rhythms was witnessed vis‐à‐vis different treatment groups. The testosterone peaks always occurred at the same time irrespective of breeding status of the bird, but with significant variation in its amplitude (high in hypergonadal and low in hypogonadal condition). It is suggested that administration of 5‐HTP and L‐DOPA at specific time interval and variation in pineal functions that modulate reproductive responses also alter the circadian pattern (acrophase and amplitude) of hypothalamic serotonin and dopamine, maintaining a specific phase relation between these cycles and breeding status. These findings strengthen our previous reports that a specific circadian phase relation of serotonergic and dopaminergic oscillations regulates reproduction. The present study strongly supports interdependence and specific relation of the two systems (gonadal activity and circadian pattern/phase relation of neural oscillation) in both natural and experimentally simulated conditions.     

Specific neural phase relation of serotonin and dopamine modulate the testicular activity in Japanese quail

Specific phase relation of serotonin and dopamine modulate the hypothalamo–hypophyseal–gonadal axis as well as photosexual responses in Japanese quail, but the effect of these specific phase relations on testicular activity and steroidogenesis is not yet been investigated. We hypothesized that temporal phase relation induced alteration in local testicular gonadotropin-releasing hormone (GnRH)–Gonadotropin-inhibitory hormone (GnIH) and their receptor system may modulate the testicular activity and steroidogenesis through local (paracrine and autocrine) action. To validate this hypothesis, we have checked the alterations in the expression of gonadotropin-releasing hormone receptor (GnRH-R), gonadotropin-inhibitory hormone receptor (GnIH-R) messenger RNA (mRNA), growth hormone receptor (GH-R), proliferating cell nuclear antigen (PCNA), cell communication and gap junctional proteins (14-3-3 and connexin-43 [Cnx-43]), steroidogenic factor-1 (SF-1), steroidogenic acute regulatory (StAR) protein, steroidogenic enzyme (3β-hydroxysteroid dehydrogenase [3β-HSD]) in testis as well as androgen receptor (AR) in testis and epididymis of control, 8-, and 12-hr quail. Experimental findings clearly indicate the increased expression of GnIH-R mRNA and suppression of GnRH-R, GH-R, PCNA, 14-3-3, Cnx-43, SF-1, StAR, 3β-HSD in testis as well as AR in testis and epididymis in 8-hr quail, while 12-hr quail exhibited the opposite results that is significantly decreased expression of GnIH-R mRNA and increased expression of GnRH-R, GH-R, PCNA, 14-3-3, Cnx-43, SF-1, StAR, 3β-HSD in testis as well as AR in testis and epididymis. The significantly increased intratesticular testosterone has been observed in the 12-hr quail while, 8-hr quail showed opposite result. Hence, it can be concluded that 12-hr quail showed significantly increased testicular activity and steroidogenesis while opposite pattern was observed in 8-hr quail.     

Role of nitric oxide in ovarian follicular development and egg production in Japanese quail (Coturnix coturnix japonica)

Role of nitric oxide (NO) in regulating the reproductive functions at hypothalamo-hypophysealovarian axis in Japanese quail was studied. In first experiment, metabolites of NO, i.e. nitrite and nitrate (NO2 and NO3) were estimated together in hypothalamus, serum and ovarian follicles of good and poor layers. In the second experiment, different NO modulators such as L-arginine (L-Arg), sodium nitroprusside (SNP) and N(G)-nitro-L-arginine methyl ester, HCl (L-NAME) were administered to the birds. In the first experiment, significantly higher (P < 0.01) NO2 and NO3 levels in serum, hypothalamus and largest (F1) ovarian follicles were observed in good layers as compared to poor layers. Higher (P < 0.05) NO2 and NO3 concentration was observed in F1 follicles than smaller follicles (F2) only in good layers. The NO2 and NO3 concentration was significantly reduced (P < 0.05) in post ovulatory follicles (POFs) in comparison to F1 and F2 follicles. In the second experiment, the serum NO2 and NO3 concentrations were higher (P < 0.05) in the SNP, lower (P < 0.05) in the L-Name group and unchanged in the L-Arg treated group in comparison to control group. compared to control, L-Arg and SNP increased (P < 0.05) the hypothalamic NO2 and NO3 concentration where as L-NAME reduced (P < 0.05) these levels. The NO2 and NO3 concentration was increased (P < 0.05) as the follicle size increased and it was significantly reduced (P < 0.05) in POFs. The higher (P < 0.05) follicular NO2 and NO3 concentration was observed in L-Arg group in comparison to control group. Egg production was also found to be higher (P < 0.05) in L-Arg group whereas it was not different (P > 0.05) in SNP and L-NAME treated groups. The yolk weight and yolk to albumin ratio was reduced (P < 0.05) in L-NAME group in comparison to control group. It may be concluded from the present study that NO plays a key role in regulating follicular development, ovulatory mechanisms and egg production in Japanese quail.     

Reproductive phase dependent circadian variation in hypothalamic concentration of serotonin, dopamine and peripheral thyroxine levels in Japanese Quail following 5-HTP and L-DOPA administration at specific time intervals

Temporal phase relations of circadian hypothalamic neurotransmitters are reported to regulate seasonal reproduction in some avian species. Present experiments were designed to study circadian variation in the hypothalamic concentration of neurotransmitters (serotonin and dopamine) and the plasma thyroxine level in sexually active (long day) and inactive (short day) Japanese Quail. A significant circadian cycle was noted in the hypothalamic content of both serotonin and dopamine, but with different patterns. In breeding Quail, peak activity of serotonin and dopamine was noted at 10.00 A.M. and 10.00 P.M. respectively i.e. at the interval of 12 hours. However, during sexually quiescent condition, peaks of both neurotransmitters occurred at 2.00 P.M. i.e. having a 0-hour temporal relationship. During the breeding phase, the plasma thyroxine level showed a biphasic pattern with two circadian peaks at 10.00 A.M. and 10.00 P.M. whereas in the non-breeding condition a single peak was observed at 10.00 A.M. In the second experiment, to study the effect of temporal synergism of neurotransmitter precursor drugs on circadian cycles, two groups of Quail were administered daily with serotonin precursor 5-HTP (5-hydroxytryptophan) and dopamine precursor L-DOPA (L-dihydroxyphenylalanine) (5 mg/100 g body weight) 12 hour (12-hr) and 8 hour (8-hr) apart over a period of 11 days under continuous conditions of light and then transferred to long day length for 15 days when the experiment was terminated. When compared to controls, the 12-hr condition induced breeding while the 8-hr condition led to a non-breeding condition. The circadian pattern of serotonin levels of control and 12-hr Quail was similar to that of a normal sexually active bird, while that of the 8-hr Quail showed the pattern of a sexually inactive bird. The plasma thyroxine level exhibited a biphasic pattern in 12-hr Quail, which was similar to a normally breeding bird, whereas unlike sexually inactive birds, the thyroxine concentration in 8-hr Quail was relatively low and did not show significant cyclicity. Interestingly, the plasma testosterone level of 12-hr Quail followed a more or less similar pattern with peak activities coinciding with that of thyroxine i.e. biphasic in the sexually active condition (12-hr and control) but a single peak in the quiescent (8-hr) condition. These findings suggest that the temporal phase relation of circadian serotonergic and dopaminergic oscillator varies as a function of reproductive status of the bird, and breeding/non-breeding conditions may be induced experimentally by changing the phase relation of these oscillations.     

Influence of different photoperiods on development of gonad, cloacal gland and circulating thyroid hormones in male Japanese quail Coturnix coturnix japonica.

One day old chicks of Japanese quail were exposed to different photoperiods (LD, 8:16, 13.5:10.5, 16:8 and LL) and observations (testes weight, cloacal gland size, body weight and circulating thyroxine and triiodothyronine) were taken at the age of 3, 5, 7, 9 and 16 weeks. Results indicate that immediate reproductive development occurred in birds exposed to long photoperiods (greater than 12 hr). Growth under LD 8:16, was not apparent till 7th week and by 16 weeks, degree of gonadal development was similar in all the birds, irrespective of photoperiodic treatment. Whereas body weight of the intermediate and long day (LD 13.5:10.5, 16:8 and LL) treated birds increased upto 5th week and remained constant thereafter. But the chicks maintained under short day length (LD 8:16), showed spontaneous increase till the end of the study and birds were much heavier compared to all other groups. Plasma T4 concentration increased with increasing age till 9th week and remained unaltered thereafter. On the other hand T3 level did not change till 7th week followed by a decline. It is suggested that the initiation and degree of gonadal growth in quail depends on the availability of daily photoperiod, until the achievement of full breeding condition. Peak level of T4 observed in 9 week old birds may be involved in the development of photorefractoriness at that age.     

Role of nitric oxide dependence on nitric oxide synthase-like activity in the water stress signaling of maize seedling

Nitric oxide (NO) has been known as an important signal in plant antioxidative defense but its production and roles in water stress are less known. The present study investigated whether NO dependence on a NO synthase-lika (NOS) activity is involved in the signaling of drought-induced protective responses in maize seedlings. NOS activity, rate of NO release and drought responses were analyzed when NO donor sodium nitroprusside (SNP), NO scavenger c-PTIO (2-(4-carboxyphenyl)-4,4,5,5-tetramathylimidazoline-1-oxyl-3-oxide) and NOS inhibitor L-NAME (NG-nitro-L-arginine methyl ester) were applied to both detached maize leaves and whole plants. Both NOS activity and the rate of NO release increased substantially under dehydration stress. The high NOS activity induced by c-PTIO as NO scavenger and NO accumulation Inhibited by NOS inhibitor L-NAME In dehydration-treated maize seedlings Indicated that most NO production under water deficit stress may be generated from NOS-like activity. After dehydration stress for 3 h, detached maize leaves pretreated with NO donor SNP maintained more water content than that of control leaves pretreated with water. This result was consistent with the decrease in the transpiration rate of SNP-treated leaves subjected to drought treatment for 3 h. Membrane permeability, a cell injury index, was lower in SNP-trested maize leaves under dehydration stress for 4 h when compared with the control leaves. Also, superoxide dismutsse (SOD) activity of SNP combined drought treatment maize leaves was higher than that of drought treatment alone, indicating that exogenous NO treatment alleviated the water loss and oxidative damage of maize leaves under water deficit stress. When c-PTIO as a specific NO scavenger was applied, the effects of applied SNP were overridden. Treatment with L-NAME on leaves also led to higher membrane permeability, higher transpiration rate and lower SOD activities than those of control leaves, indicating that NOS-like activity was involved in the antioxidative defense under water stress. These results suggested that NO dependence on NOS-like activity serves as a signaling component in the induction of protective responses and is associated with drought tolerance in maize seedlings.     

NO和NADPH-黄递酶在绿豆下胚轴不定根发生和发育过程中的变化

研究了一氧化氮(NO)供体普钠(SNP)、一氧化氮清除剂C-PTIO和一氧化氮合酶(NOS)抑制L-NAME对绿豆(Vigna radiataL.)下胚轴插条生根的影响.并对不定根生期间手条基部NO 和NADPH-黄递酶的时空变化进行了检测.所试浓度SNP均明显促进下胚轴不根发生.分别插条切取后24h和36h于其基部维管束之间检测到NADPH-黄递酶(NOS标记酶)阳性反应和NO荧光,根原基也于48h在相同位置出现,并于60h进一步伸长.48~60h期间,NADPH、黄递的阳性反应及NO荧光有增强趋势,并主要分布在不定根分生组织中.L-NAME既减弱NADPH-黄递酶的阳性反应和NO荧光,也延缓不不定根发生;而c-PTIO对NO荧光及不定根生均有抑制作用.上述结果证明:NO在不定根发生及发育过程中有重要作用,而且此过程中的NO很可能由类似的NOS催化产生.     

Characterization of Recombinant Human Aromatic l-Amino Acid Decarboxylase Expressed in COS Cells

The expression vector containing the full-length cDNA of human aromatic L-amino acid decarboxylase (EC 4.1.1.28) was transfected in COS cells by a modified calcium phosphate coprecipitation method. The cells transfected with plasmids that had a true direction of the cDNA gave a major immunoreactive band at 50 kDa. This expressed enzyme catalyzed the decarboxylation of L-3,4-dihydroxyphenylalanine (L-DOPA), L-5-hydroxytryptophan (L-5-HTP) and L-threo-3,4-dihydroxyphenylserine. The optimal pH of the enzyme activity with L-DOPA as a substrate was 6.5, whereas the enzyme had a broad pH optimum when L-5-HTP was used as a substrate. Addition of pyridoxal phosphate to the incubation mixture greatly enhanced the activity for both L-DOPA and L-5-HTP.     

Requirement of nitric oxide for murine oocyte maturation, embryo development, and trophoblast outgrowth in vitro

We investigated the extent to which NO participates in the developmental competence (oocyte maturation, fertilization and embryo development to blastocyst) using an in vitro culture system adding sodium nitroprusside (SNP), NO donor, and NOS inhibitor (N-omega-nitro-L-arginine methyl ester, L-NAME). We also assessed the effects of NO/NOS system on blastocyst implantation using an in vitro trophoblast outgrowth assay. The treatment of low concentrations of SNP (10(-7) M) significantly stimulated meiotic maturation to metaphase II stages in cumulus enclosed oocytes. In contrast, 10(-3) and 10(-5) M L-NAME demonstrated a significant suppression in resumption of meiosis. This inhibition was reversed by the addition of SNP. No development beyond the four-cell stage was observed by the addition of high concentration of SNP (10(-3) M). Inhibition of embryo development, especially the conversion of morulae to blastocysts, was also observed in the treatment of lower doses of SNP (10(-5) and 10(-7) M). Similarly, inhibition of NO by NOS inhibitor resulted in the dose-dependent inhibition of embryo development and hatching rates, but the concomitant addition of SNP with L-NAME reversed the inhibitory effect by each SNP or L-NAME treatment. Furthermore, low concentration of SNP (10(-7) M) but not high concentration of SNP (10(-3) M) significantly stimulated trophoblast outgrowth, whereas the addition of L-NAME suppressed the spreading of blastocysts in a dose-dependent manner. These results suggest that NO may have crucial roles in oocyte maturation and embryogenesis including the process of implantation. The observed differences in required amount of NO and the sensitivity to cytotoxicity of NO in each developmental stage embryos may also suggest that NO/NOS system is tightly regulated in developmental stage specific manner.     

NO和NADPH-黄递酶在绿豆下胚轴不定根发生和发育过程中的变化(英文)

研究了一氧化氮(NO)供体硝普钠(SNP)、一氧化氮清除剂c-PTIO和一氧化氮合酶(NOS)抑制剂L-NAME对绿豆(Vigna radiata L.)下胚轴插条生根的影响,并对不定根发生期间插条基部NO和NADPH-黄递酶的时空变化进行了检测。所试浓度SNP均明显促进下胚轴不定根发生。分别在插条切取后24 h和36 h于其基部维管束之间检测到NADPH-黄递酶(NOS标记酶)阳性反应和NO荧光,根原基也于48 h在相同位置出现,并于60 h进一步伸长。48-60h期间,NADPH-黄递酶的阳性反应及NO荧光有增强趋势,并主要分布在不定根分生组织中。L-NAME既减弱NADPH-黄递酶的阳性反应和NO荧光,也延缓不定根发生;而c-PTIO对NO荧光及不定根发生均有抑制作用。上述结果证明:NO在不定根发生及发育过程中有重要作用,而且此过程中的NO很可能由类似的NOS催化产生。     

Presence of endogenous inhibitor of aromatic L-amino acid decarboxylase in monkey serum

Summary In the course of our studies on the developmental changes of aromatic L-amino acid decarboxylase (AADC) in the serum of Japanese monkeys (Macaca fuscata fuscata), we found the presence of an endogenous inhibitor of AADC in all stages of monkey life. This inhibitor inhibited the serum enzyme activity completely with L-5-hydroxytryptophan (L-5-HTP) as substrate, while the activity was partially inhibited with L-DOPA as substrate. The inhibitor was non-dialyzable, but it could be removed from the monkey serum by DEAE-Sephacel chromatography. After this treatment AADC activities could be detected in the monkey serum by using both L-DOPA and L-5-HTP as substrates. Moreover, the total activity for L-DOPA was augmented by 3-fold in the serum after the removal of the inhibitor. Serum AADC was partially purified from monkey and compared with that of rat using both L-DOPA and L-5-HTP as substrates, but the ratio of the activities for the two substrates did not change significantly in each fraction during purification from either monkey or rat serum.On leave from the University of Rajshahi, Rajshahi, Bangladesh.     

一氧化氮合酶抑制剂L-NAME对大鼠脑缺血耐受诱导的影响

采用大鼠四血管闭塞全脑缺血耐受模型和脑组织切片形态学方法,观察应用一氧化氮合酶(NOS)抑制剂L—NAME对大鼠海马CAl区脑缺血耐受(BIT)诱导的影响,在整体水平探讨一氧化氮(NO)在BIT诱导中的作用。54只Wistar大鼠凝闭双侧推动脉后分为6组：(1)假手术组(n＝6);分离双侧颈总动脉,但不阻断脑血流;(2)损伤性缺血组(n＝6)：全脑缺血10min;(3)预缺血 损伤性缺血组(n＝6)：脑缺血预处理(CIP)3min,再灌注72h后行全脑缺血10min;(4)L—NAME组;分别于CIP前1h和后1、12及36h腹腔注射L—NAME(5mg／kg),每个时间点6只动物,其余步骤同预缺血 损伤性缺血组;(5)L—NAME L—精氨酸组(n＝6)：于CIP前1h腹腔注射L—NAME(5mg／kg)和L—精氨酸(300mg／kg),其它步骤同L—NAME组;(6)L—NAME 损伤性缺血组(n＝6)：于腹腔注射L—NAME(5mg／kg)72h后行全脑缺血10min。实验结果表明,(1)单纯10min全脑缺血可使海马CAl区组织学分级增加(表明损伤加重),神经元密度降低(P＜0．01);(2)预缺血 损伤性缺血组的海马CAl区组织学分级、神经元密度与假手术组相比,无显著性差别(P＞0．05);(3)L—NAME组中,应用L—NAME后海马CAl区组织学分级增加,神经元密度降低,与预缺血 损伤性缺血组相比有显著性差异(P＜0．05),表明L—NAME可阻断CIP对神经元的保护作用;(4)L—NAME L—精氨酸组与L—NAME组相比,海马CAl区组织损伤明显减轻(P＜0．05),但与预缺血 损伤性缺血组相比仍有显著性差别(P＜0．05),提示L-精氨酸可部分逆转L—NAME的作用;(5)L—NAME 损伤性缺血组的组织学表现与损伤性缺血组相同(P＞0．05)。这些结果表明,在整体情况下N0参与BIT的诱导。与CIP前1h及后1、12h给予L—NAME组相比,CIP后36h给予L—NAME对CIP保护作用的阻断效应明显减弱,提示N0在CIP后较早阶段即开始参与BIT的诱导。     

Depression of plasma luteinizing hormone concentration in quail by the anticholinesterase insecticide parathion

To examine the effects of parathion on basal plasma luteinizing hormone (LH) concentration, male Japanese quail (Coturnix japonica) were orally intubated with 0, 5 or 10 mg/kg parathion and sacrificed after 4, 8 and 24 hr. At the 5 mg/kg dose, plasma LH levels were reduced at 4 and 8 hr, but returned to control values by 24 hr. Brain acetylcholinesterase activity was substantially reduced by 10 mg/kg parathion (52, 75 and 37% inhibition at 4, 8 and 24 hr, respectively) and plasma LH concentration remained depressed through the 24-hr period. These findings suggest that the organophosphorus insecticide parathion may alter plasma LH concentration in a manner which might impair reproductive activity, and provide indirect evidence for a cholinergic component in the regulation of LH secretion in quail.     

Sex differences in modulating blood brain barrier permeability by NO in pentylenetetrazol-induced epileptic seizures

Susceptibility to epilepsy as well as BBB dysfunction in some pathological conditions varies depending on sex difference. It has recently been shown that systemically given NO donor and antagonists modify the nature of seizures induced by PTZ (pentylenetetrazol) differently in male and female rats. This study investigates the role of NO on BBB permeability in PTZ seizures with sex differences using NO donor, sodium nitroprusside (SNP), and NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME). Nitrite+nitrate levels as indices of NO generation in the brain were also assessed. L-NAME prolonged seizure latency in male rats, seizure intensity and seizure duration were lessened. L-NAME depicted opposite effects in seizure nature in female rats. SNP prolonged seizure latency, while seizure intensity and duration were lessened only in female rats. L-NAME in male rats increased L-NAME use in female rats (not in male rats) which resulted in a more leaky BBB especially in midbrain, thalamus, hippocampus, corpus striatum and cerebellum whereas SNP use in male rats and not in female rats resulted in pronounced BBB opening in all brain regions studied than PTZ per se. L-NAME while decreasing nitrite+nitrate levels in male rat brains, acted in an opposite fashion in females. SNP use depicted an inverse picture with respect to L-NAME, with an opposite action in different sexes. This study reveals that NO effect on BBB in PTZ-induced seizures depends unequivocally on sex difference. The sex-dependent action of NO in seizures and in CNS pathologies warrants further investigation.     

IV. Determination of aromatic L-amino acid decarboxylase in serum of various animals by high-performance liquid chromatography with electrochemical detection

This paper describes the distribution of aromatic L-amino acid decarboxylase (AADC), using both L-DOPA and L-5-hydroxytryptophan (L-5-HTP) as substrates, in serum of various animals. The ratio of the activities of the enzyme towards both substrates was also determined in the same serum. AADC activity was discovered in serum using our new and highly sensitive assay for AADC activity by high-performance liquid chromatography (HPLC) with electrochemical detection (ED) with L-DOPA and L-5-HTP as substrates. It was found that among the species used, guinea pig serum had the highest activity, and we made a systematic study on guinea pig serum AADC.     

Purification and characterization of L-DOPA decarboxylase from human kidney

L-DOPA decarboxylase has been purified to homogeneity from post mortem removed human kidneys. Homogeneity was examined by polyacrylamide gel electrophoresis (PAGE) analysis both in the presence and absence of SDS. The enzyme has a molecular weight of 100,000 daltons estimated by gel filtration and 50,000 daltons determined after SDS-PAGE. Human L-DOPA decarboxylase therefore is a dimer. Polyclonal antibodies produced against human L-DOPA decarboxylase react with the 50,000 daltons enzyme subunit after immuno-blotting and also precipitates enzyme activity. Activity against L-DOPA is partially inhibited by 5-hydroxytryptophan (5-HTP). The effect of various cations on L-DOPA decarboxylase activity has also been tested.     

The Interactions of Nitric Oxide and Acetylcholine on Penicillin-Induced Epilepsy in Rats

The aim of this study was to investigate the interaction between nitric oxide (NO) and acetylcholine (ACh) in penicillin-induced experimental epilepsy. Adult male Wistar rats weighing 220 ± 35 g were used in the experiments. The epileptiform activity was induced by microinjection of penicillin (200 IU/1 μl) into the left sensorymotor cortex. Electrocorticogram was recorded by using Ag/AgCl ball electrodes. Sodium nitroprusside (SNP), a NO donor, given intracortically 30 min after penicillin significantly reduced the spike frequency whereas ACh increased the epileptiform activity for 5 min. Atropine, an antagonist for muscarinic receptors, was given intracortically 30 min after penicillin and did not significantly affect epileptiform activity for 30 min. SNP given after atropine significantly suppressed the epileptiform activity. ACh given 10 min after Nω-nitro-L: -arginine methyl ester (L-NAME), a nonspecific nitric oxide synthase inhibitor, did not have a significant effect on spike frequency. When ACh and SNP were administered together, penicillin induced epileptiform activity and spike frequency were significantly suppressed from the 10th minute onwards. It can be concluded that ACh increases the penicillin-induced epileptiform activity while co-administration of ACh and SNP produces a potent anticonvulsant effect as compared to SNP alone.