Inhibition of endogenous hydrogen sulfide biosynthesis enhances the anti-cancer effect of 3,3'-diindolylmethane in human gastric cancer cells.

AIMS: 3,3'-Diindolylmethane (DIM) has limited anti-cancer effects in gastric cancer. Hydrogen sulfide (H2S) plays an important role in the tumor development and therapy, cystathionine-ß-synthase (CBS) and cystathionine-γ-lyase (CSE), two key endogenous H2S biosynthesis enzymes, can affect endogenous H2S levels and alter cancer treatment. Our main objective was to investigate whether the aminooxyacetic acid (AOAA) and DL-Propargylglycine (PAG), two specific inhibitors of CBS and CSE, could assist DIM to exert a stronger anti-cancer effects in gastric cancer BGC-823 and SGC-7901 cells. MATERIALS AND METHODS: Cell proliferation was assayed by MTT and cell colony-forming assay. Apoptosis and migration were detected by Hoechst staining and scratch test respectively. Western blot was used to evaluate the expression of proteins related to proliferation, apoptosis and migration. KEY FINDINGS: Combination of AOAA or PAG with DIM synergistically inhibited proliferation and migration, increased apoptosis in gastric cancer cells. The p38-p53 axis was also further activated by the combination of AOAA or PAG with DIM. Exogenous H2S from sodium hydrosulfide, attenuated the efficacy of DIM in cancer cells by reducing the activation level of p38-p53 axis. Taken together, AOAA or PAG inhibited the expression of endogenous H2S biosynthesis enzymes and effectively enhanced susceptibility of gastric cancer to DIM through activating p38-p53 axis. SIGNIFICANCE: The current study highlight more precise requirements for the clinical application of sulfur-containing anti-cancer drugs, and open a new way to enhance the sensitivity of DIM in chemotherapy of gastric cancer.

Central administration of aminooxyacetate, an inhibitor of H2S production, affects thermoregulatory but not cardiovascular and ventilatory responses to hypercapnia in spontaneously hypertensive rats.

Hydrogen sulfide (H2S) is classically known for its toxic effects. More recently H2S has been documented as a neuromodulator. Here we investigated the central effects of aminooxyacetate (AOA; inhibitor of the H2S-synthesizing enzyme cystathionine ß-synthase, CBS) on cardiovascular, respiratory and thermoregulatory responses to hypercapnia in spontaneously hypertensive rats (SHR). To attain this goal we measured mean arterial pressure (MAP), heart rate (HR), ventilation (VE), and deep body temperature (Tb) of SHR and (normotensive) Wistar Kyoto (WKY) rats before and after microinjection of AOA (9 nmol/µL) or saline into the fourth ventricle immediately followed by 30-min hypercapnia exposure (7% inspired CO2). In saline-treated WKY rats, hypercapnia caused an increase in MAP accompanied by bradycardia, an increase in VE, and a drop in Tb. In AOA-treated WKY rats exposed to hypercapnia, the drug did not affect the increased MAP, potentiated the bradycardic response, attenuated the increased VE, and potentiated the drop in Tb. In saline-treated SHR, in comparison to the saline-treated WKY rats, hypercapnia elicited a minor, shorter-lasting increase in MAP with no changes in HR, evoked a greater increase in VE, and did not induce a drop in Tb. In AOA-treated SHR exposed to hypercapnia, the drug did not change the hypercapnia-induced cardiovascular and ventilatory responses while permitted a drop in Tb. Our findings indicate that AOA, an inhibitor of H2S production, modulates cardiorespiratory and thermoregulatory responses to hypercapnia in normotensive rats, whereas hypertension development in SHR is accompanied by suppression of the AOA effect on the cardiovascular and respiratory responses.

Effects of aminooxyacetic acid on hippocampal mitochondria in rats with chronic alcoholism: the analysis of learning and memory-related genes.

The incidence of chronic alcoholism leading to central and peripheral nervous system damage has been increasing year-to-year. The purpose of this study is to explore the effects of aminooxyacetic acid on hippocampus mitochondria in rats with chronic alcoholism and analyze learning and memory-related genes. Sixty male Sprague Dawley rats were randomly divided into three groups. Except for the control group, each group was fed with the water containing (v/v) 6% alcohol for 28 days. After 14 days, rats in the treatment group were intraperitoneally injected daily for 14 days with aminooxyacetic acid. High throughput sequencing was combined and tested for learning and memory abilities, Hydrogen sulfide content, catalase activity in mitochondria, and the expression of F-actin in the hippocampus of the rats in each group. Compared with the control group, the learning and memory abilities of rats with chronic alcoholism were significantly impaired, mitochondria contained vacuoles, hydrogen sulfide increased, but catalase activity and F-actin content were significantly decreased, After treatment with aminooxyacetic acid, mitochondrial morphology improved, hydrogen sulfide content was decreased, while catalase activity and F-actin expression of in hippocampus were increased. This indicates that aminooxyacetic acid may improve learning and memory in rats with chronic alcoholism, and the mechanism is related to decreased hydrogen sulfide content and an increase of both catalase activity and F-actin level in the hippocampus, thereby reducing the damage of alcohol to mitochondria and neurons.

Involvement of endogenous central hydrogen sulfide (H2S) in hypoxia-induced hypothermia in spontaneously hypertensive rats.

Spontaneously hypertensive rats (SHR) display autonomic imbalance and abnormal body temperature (Tb) adjustments. Hydrogen sulfide (H2S) modulates hypoxia-induced hypothermia, but its role in SHR thermoregulation is unknown. We tested the hypothesis that SHR display peculiar thermoregulatory response to hypoxia and that endogenous H2S overproduced in the caudal nucleus of the solitary tract (NTS) of SHR modulates this response. SHR and Wistar rats were microinjected into the fourth ventricle with aminooxyacetate (AOA, H2S-synthezing enzyme inhibitor) or sodium sulfide (Na2S, H2S donor) and exposed to normoxia (21% inspired O2) or hypoxia (10% inspired O2, 30 min). Tb was continuously measured, and H2S production rate was assessed in caudal NTS homogenates. In both groups, AOA, Na2S, or saline (i.e., control; 1 µL) did not affect euthermia. Hypoxia caused similar decreases in Tb in both groups. AOA presented a longer latency to potentiate hypoxic hypothermia in SHR. Caudal NTS H2S production rate was higher in SHR. We suggest that increased bioavailability of H2S in the caudal NTS of SHR enables the adequate modulation of excitability of peripheral chemoreceptor-activated NTS neurons that ultimately induce suppression of brown adipose tissue thermogenesis, thus accounting for the normal hypoxic hypothermia.

Endogenous preoptic hydrogen sulphide attenuates hypoxia-induced hyperventilation.

AIM: We hypothesized that hydrogen sulphide (H2 S), acting specifically in the anteroventral preoptic region (AVPO - an important integrating site of thermal and cardiorespiratory responses to hypoxia in which H2 S synthesis has been shown to be increased under hypoxic conditions), modulates the hypoxic ventilatory response. METHODS: To test this hypothesis, we measured pulmonary ventilation (V˙E) and deep body temperature of rats before and after intracerebroventricular (icv) or intra-AVPO microinjection of aminooxyacetate (AOA; CBS inhibitor) or Na2 S (H2 S donor) followed by 60 min of hypoxia exposure (7% O2 ). Furthermore, we assessed the AVPO levels of H2 S of rats exposed to hypoxia. Control rats were kept under normoxia. RESULTS: Microinjection of vehicle, AOA or Na2 S did not change V˙E under normoxic conditions. Hypoxia caused an increase in ventilation, which was potentiated by microinjection of AOA because of a further augmented tidal volume. Conversely, treatment with Na2 S significantly attenuated this response. The in vivo H2 S data indicated that during hypoxia the lower the deep body temperature the smaller the degree of hyperventilation. Under hypoxia, H2 S production was found to be increased in the AVPO, indicating that its production is responsive to hypoxia. The CBS inhibitor attenuated the hypoxia-induced increase in the H2 S synthesis, suggesting an endogenous synthesis of the gas. CONCLUSION: These data provide solid evidence that AVPO H2 S production is stimulated by hypoxia, and this gaseous messenger exerts an inhibitory modulation of the hypoxic ventilatory response. It is probable that the H2 S modulation of hypoxia-induced hyperventilation is at least in part in proportion to metabolism.

N-Acetylcarnosine and histidyl-hydrazide are potent agents for multitargeted ophthalmic therapy of senile cataracts and diabetic ocular complications.

AIMS: In human diabetes, the deleterious effects of chronic hyperglycemia are the result of excessive nonenzymatic modification of proteins and phospholipids by glucose and its by-products leading to the formation of irreversible oxidized, aromatic, and fluorescent ligands known as advanced glycation end products. This glycation process has been associated with deleterious health effects. The present invention provides the potent inhibitors of protein glycation and AGEs formation, which are particularly advantageous for eyedrop delivery in the prevention and treatment of diabetes- and age-related pathologies. MAIN METHODS AND KEY FINDINGS: We proposed a deglycation system involving removal, by transglycation of sugar or aldehyde moieties from the Schiff bases by ophthalmic aldehyde scavenger L-carnosine derived from its ocular bioactivating sustained release prodrug 1% N-acetylcarnosine (NAC) lubricant eyedrops containing a mucoadhesive cellulose compound combined with corneal absorption promoters in drug delivery system. Carnosine analogs bearing the histidyl-hydrazide moiety were synthesized and patented in ophthalmic formulations with NAC bioactivating prodrug to moderate the enzymatic hydrolysis of a dipeptide by carnosinase (inhibited by a nonhydrolyzable substrate analog so that this keeps steadier levels of the drug active principle in the aqueous humor). Leucyl-histidylhydrazide peptidomimetic demonstrated the transglycation activity more pronounced than L-carnosine accounting for the ability of either molecule to reverse pre-existing, glycation-induced, cross-linking, and checking the nonenzymatic glycation cascade in the ophthalmic pathologies. The ophthalmic drug N-acetylcarnosine eye drop formulation with sustained time- release and increased absorption of L-carnosine in the aqueous humor (a prolonged effective dose) showed follow-up treatment efficacy for age-related cataracts for enrolled patients into the randomized double blind placebo controlled crossover clinical trial, and in over 50250 various cohort patients, was demonstrated to have an efficacy, safety and good tolerability for prevention and treatment of visual impairment in the older population data base. SIGNIFICANCE: The bioactivating antioxidant NAC and histidyl-hydrazide are potent agents with the pleiotropic effects for ophthalmic therapy of senile cataracts and diabetic ocular complications.

Kynurenate production by cultured human astrocytes.

In the rodent brain, astrocytes are known to be the primary source of kynurenate (KYNA), an endogenous antagonist of both the glycine(B) and the alpha7 nicotinic acetylcholine receptor. In the present study, primary human astrocytes were used to examine the characteristics and regulation of de novo KYNA synthesis in vitro. To this end, cells were exposed to KYNA's bioprecursor L-kynurenine, and newly formed KYNA was recovered from the extracellular milieu. The production of KYNA was stereospecific and rose with increasing L-kynurenine concentrations, reaching a plateau in the high microM range. In an analogous experiment, astrocytes also readily produced and liberated the potent, specific glycine(B) receptor antagonist 7-chlorokynurenate from L-4-chlorokynurenine. KYNA synthesis was dose-dependently reduced by L-leucine or L-phenylalanine, two amino acids that compete with L-kynurenine for cellular uptake, and by aminooxyacetate, a non-specific aminotransferase inhibitor. In contrast, KYNA formation was stimulated by 5 mM pyruvate or oxaloacetate, which act as co-substrates of the transamination reaction. Aglycemic or depolarizing (50 mM KCl or 100 microM veratridine) conditions had no effect on KYNA synthesis. Subsequent studies using tissue homogenate showed that both known cerebral kynurenine aminotransferases (KAT I and KAT II) are present in astrocytes, but that KAT II appears to be singularly responsible for KYNA formation under physiological conditions. Taken together with previous results, these data suggest that very similar mechanisms control KYNA synthesis in the rodent and in the human brain. These regulatory events are likely to influence the neuromodulatory effects of astrocyte-derived KYNA in the normal and diseased human brain.

[Effect prolonged GABAergic activation on the pubertal development of female rats]. / Efecto de la activación GABAérgica sobre la maduración sexual en ratas hembras peripuberales.

We evaluated, in immature female rats, the effect of the GABAergic system on the reproductive axis and on pubertal development. Initially, using a prolonged treatment with aminooxyacetic acid (AOAA), increasing hypothalamic GABA (p < 0.002), and decreasing GnRH and glutamate content (p < 0.05 and < 0.02). Treated rats showed diminished serum LH (p < 0.05) and estradiol (p < 0.005) levels. Vaginal opening occurred at 30.8 +/- 0.6 days in controls, and at 36.7 +/- 0.98 days in AOAA-treated rats. Acute treatment with AOAA resulted in a decreased GnRH and glutamate output, and in an increased taurine release from superfused hypothalamic fragments. This effect was mimicked by the GABA-A and GABA-B agonists. The activation of the GABAergic system during postnatal days 23-29 significantly restrains the hypothalamo-pituitary-ovaric axis and delays the onset of puberty. The existence of a physiological cross-talk between excitatory and inhibitory amino acid neurotransmitters regulating GnRH release during the onset of puberty is postulated.

Identification of the 1-cyano-3,4-epithiobutane-derived urinary mercapturic acid N-acetyl-S-(4-cyano-2-thio-1-butyl)-cysteine in male Fischer 344 rats.

1-Cyano-3,4-epithiobutane (CEB), a naturally occurring nitrile derived from cruciferous plants, causes nephrotoxicity in male Fischer 344 rats. Nephrotoxicity induced by CEB is dependent on glutathione (GSH) conjugation and bioactivation. Conjugation with GSH and subsequent metabolism leads to the formation of specific urinary metabolites. The objectives of the present study were to identify CEB-derived urinary metabolites and quantify urinary non-protein thiols and thioethers in male Fischer 344 rats. Animals received 125 mg kg(-1) of CEB alone or following pretreatment with one of three selective inhibitors of GSH metabolism: acivicin, probenecid or aminooxyacetic acid. Total non-protein urinary thiol and urinary thioether concentrations were elevated in all treated groups at 12 and 24 h; however, elevations in non-protein thiols were not significantly greater in rats administered CEB alone as compared to negative controls. A single predominant urinary metabolite was identified as the CEB-derived mercapturic acid N-acetyl-S-(4-cyano-thio-1-butyl)-cysteine. Evidence for other CEB-derived metabolites was also demonstrated. These findings represent the identification of a unique compound and provide further evidence for the importance of GSH conjugation as a significant pathway in CEB metabolism.

Efecto de la activacion gabaergica sobre la maduracion sexual en ratas hembras peripuberales / Effect prologed gabaergic activation on the pubertal development of female rats

Evaluamos 1) efecto del tratamiento prolongado (días 23-29 postnatales) con ácido aminooxiacético (AAOA) sobre el desarrollo puberal en ratas hembra; este tratamiento aumentó el contenido de GABA (p< 0.002), disminuyendo el de GnRH y glutamato (p < 0.05 y < 0.02) en hipotálamo. La LH (p < 0.05) y el estradiol (p < 0.005) séricos cayeron. La apertura vaginal fue a los 30.8 + 0.6 días en los controles, y a los 36.7 + 0.98 días en las tratadas (p < 0.0001). 2) A los 30 días, el tratamiento agudo con AAOA redujo la liberación ex vivo de GnRH y de glutamato la de taurina. Este efecto fue similar al observado agregando al medio agonistas GABA-A y B. Conclusiones: la activación peripuberal del sistema GABAérgico frena el eje reprodutor, produciendo un retraso en el desarrollo. Esto podría atribuirse a la existencia, en esta etapa, de interrelaciones fisiológicas entre los aminoácidos que regulan la secreción de GnRH (GABA, glutamato, taurina). (AU)

Efecto de la activacion gabaergica sobre la maduracion sexual en ratas hembras peripuberales / Effect prologed gabaergic activation on the pubertal development of female rats

Evaluamos 1) efecto del tratamiento prolongado (días 23-29 postnatales) con ácido aminooxiacético (AAOA) sobre el desarrollo puberal en ratas hembra; este tratamiento aumentó el contenido de GABA (p< 0.002), disminuyendo el de GnRH y glutamato (p < 0.05 y < 0.02) en hipotálamo. La LH (p < 0.05) y el estradiol (p < 0.005) séricos cayeron. La apertura vaginal fue a los 30.8 + 0.6 días en los controles, y a los 36.7 + 0.98 días en las tratadas (p < 0.0001). 2) A los 30 días, el tratamiento agudo con AAOA redujo la liberación ex vivo de GnRH y de glutamato la de taurina. Este efecto fue similar al observado agregando al medio agonistas GABA-A y B. Conclusiones: la activación peripuberal del sistema GABAérgico frena el eje reprodutor, produciendo un retraso en el desarrollo. Esto podría atribuirse a la existencia, en esta etapa, de interrelaciones fisiológicas entre los aminoácidos que regulan la secreción de GnRH (GABA, glutamato, taurina).

Electrical stimulation activates two different sites within the guinea pig cochlea.

This study investigates whether auditory brainstem responses (ABRs) can be used to assess the functioning of electrically stimulated cochleas. Electrically evoked auditory brainstem responses (EABRs) were recorded in guinea pigs with normal hearing and guinea pigs deafened by amikacin, a powerful ototoxic antibiotic, combined with diuretic aminooxyacetic acid (AOAA). Two different types of EABRs were observed in normal animals, depending on the electrical pulse intensity applied to the round window: long-latency brainstem responses were evoked by low stimulation intensities, short-latency brainstem responses by high intensities. The absence of effect of strychnine applied intracochlearly ruled out the possibility of medial efferents being involved in these responses. Conversely, an intracochlear application of tetrodotoxin (TTX), an Na(+)-channel blocker, resulted in the disappearance of both types of responses, attesting that the sites activated by the electrical stimulation were located within the cochlea. In AOAA/ amikacin poisoned cochleas, in which most of the hair cells were missing with apparently normal ganglion neurons, the long-latency brainstem responses evoked by low intensities were completely lacking. These findings suggest that low currents applied to the round window of the guinea pig cochlea primarily activate the hair cells, the neurons being directly excited at higher intensities.

Acetaminophen nephrotoxicity in the CD-1 mouse. II. Protection by probenecid and AT-125 without diminution of renal covalent binding.

Acetaminophen (APAP) administration (600 mg/kg, ip) to 18-hr fasted, 3-month-old male CD-1 mice results in necrosis of the convoluted renal proximal tubules with a corresponding elevation of plasma urea nitrogen (BUN). Administration of the gamma-glutamyl transpeptidase inhibitor, L-(alpha S,5S)-alpha-amino-3-chloro-4,5-dihydroxy- 5-isoxazoleacetic acid (AT-125) (50 mg/kg, ip), to mice 30 min before APAP significantly diminished the APAP-induced histopathologic damage and BUN elevation. Administration of the organic-anion transport inhibitor, probenecid (150 mg/kg, ip), 30 min before APAP challenge also protected against the APAP-induced elevation of BUN and detectable histopathologic changes. By contrast, pretreatment of mice with the cysteine conjugate beta-lyase inhibitor, (aminooxy)acetic acid (100 mg/kg, ip), 1 hr before APAP did not alter nephrotoxicity. None of the pretreatments altered the APAP-induced elevation of plasma sorbitol dehydrogenase activity, nor were there any detectable changes in liver histopathology after APAP challenge. Despite the protective effects of both probenecid and AT-125 against nephrotoxicity, they did not affect either the level of immunochemically detectable covalent binding to protein or the depletion of renal glutathione at 4 hr after APAP. Thus, the protection appears independent of effects on renal APAP uptake or activation and indirectly suggests that an APAP-glutathione conjugate may contribute to the observed nephrotoxicity.

Differential neuronal vulnerability to amino-oxyacetate and quinolinate in the rat parahippocampal region.

Injection of the "indirect" excitotoxin amino-oxyacetate into the entorhinal area causes acute behavioral seizures and preferential neuronal loss in layer III of the medial entorhinal cortex in rats. We examined here whether the effects of amino-oxyacetate could be duplicated by local injections of the endogenous N-methyl-D-aspartate receptor agonist and direct excitotoxin, quinolinate. Amino-oxyacetate (685 nmol) or quinolinate (30, 45 or 60 nmol) were injected into the entorhinal cortex of rats anesthetized with choral hydrate (360 mg/kg). Separate groups of animals were co-treated with the N-methyl-D-aspartate receptor antagonist dizocilpine maleate (2 mg/kg) or given a higher dose of chloral hydrate (500 mg/kg). Rats that received amino-oxyacetate and a low anesthetic dose consistently displayed acute behavioral seizures and showed preferential loss of neurons in layer III of the medial entorhinal cortex. Animals that were given quinolinate did not display behavioral seizures, and showed preferential degeneration of neurons in layer V of the entorhinal cortex. Moreover, quinolinate-injected rats frequently exhibited neuronal loss in the superficial layers of the dorsal perirhinal cortex. The behavioral and neuropathological sequelae of amino-oxyacetate, but not quinolinate-induced neurotoxicity, were abolished by prolonged chloral hydrate anesthesia. In spite of these apparent qualitative differences between the two toxins, neurodegeneration induced by either amino-oxyacetate or quinolinate was completely prevented by dizocilpine maleate. These data suggest that a heterogeneous distribution of pharmacologically distinct N-methyl-D-aspartate receptor subtypes in the parahippocampal region may underlie the distinct neurodegenerative properties of the two toxins. Since the lesion caused by amino-oxyacetate bears remarkable similarities to neuropathological changes which have been described in this structure in temporal lobe epilepsy, further elucidation of the mechanisms of cellular toxicity of amino-oxyacetate may hold clues for the pathogenesis of this disease.

The influence of GABA-ergic system on clonidine analgesia in rats.

The effect of agonists and antagonists of GABA-ergic system on analgesia induced by clonidine was investigated in the rat. The compounds activating (muscimol, 1 mg/kg and aminooxyacetic acid; AOAA, 25 mg/kg) or inhibiting GABA-ergic receptors (bicuculline and picrotoxin, in subconvulsive doses 0.5-1 mg/kg) do not change clonidine-induced analgesia. Only the higher dose (2 mg/kg) of muscimol, bicuculline or picrotoxin affected clonidine-induced analgesia. Baclofen, contrary to other GABA agonists, markedly potentiated clonidine-induced analgesia in the dose dependent manner. These results suggest that compounds activating and inhibiting GABA A receptors (muscimol, AOAA, bicuculline and picrotoxin) have a weak influence on clonidine induced analgesia as compared to the effect of baclofen (agonist of GABA B receptors).

Aminooxyacetic acid-induced accumulation of GABA enhances clonidine hypotension.

The effect of 2-oxoglutarate aminotransferase inhibitor, aminooxyacetic acid (AOAA) on hypotension induced by clonidine in spontaneously hypertensive rats (SHR) was examined. AOAA given 15 min before clonidine significantly intensified clonidine--induced reduction of blood pressure. This effect reached its maximum within 120 min after AOAA administration, when AOAA itself had no influence on blood pressure. Our data suggest that the activation of functional state of GABAergic system by GABA accumulation potentiates clonidine hypotension.

Contrasting effects of the GABA transaminase inhibitors gamma-vinylGABA and aminooxyacetic acid on arterial pressure following injection into nucleus tractus solitarius.

The effect of injection of inhibitors of GABA-transaminase into the nucleus tractus solitarius (NTS) on arterial pressure was examined in anesthetized rats. Two typical GABA-transaminase inhibitors, gamma-vinylGABA (GVG) and aminooxyacetic acid (AOAA), increased arterial pressure when injected into the NTS, a response similar to that elicited by direct stimulation of GABA receptors in the NTS. However, with neither drug was the response consistent with the action of inhibiting GABA-T. The response elicited by GVG, despite GVG acting as an irreversible inhibitor of GABA-T, could be reproduced with subsequent injections. Still, the pressor response to GVG could be reversed by blockade of GABAB-receptors with CGP-35348, suggesting that GVG injection did increase GABA-mediated neural transmission. In contrast, the pressor response to AOAA could not be antagonized by blockade of either GABAA or GABAB receptors. These results suggest that acute inhibition of GABA-T does not necessarily result in an increase in GABA-mediated neural transmission.

The role of GABA-ergic signal in the regulation of melatonin biosynthesis in vertebrate retina.

The in vivo effects of GABA-ergic drugs on the activity of serotonin N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT), two enzymes involved in melatonin biosynthesis, were investigated in light-exposed chicken retina. The ip administration of muscimol and baclofen (direct agonists of GABA-A and GABA-B receptors, respectively), aminooxyacetic acid (an inhibitor of GABA transaminase), and nipecotic acid (an inhibitor of GABA reuptake), significantly increased the retinal NAT activity by 50-100%. Similar rises in NAT activity were observed following intraocular treatment of ether-anesthetized chickens with muscimol, baclofen and GABA. In contrast to NAT, there was no effect of the tested drugs on the retinal HIOMT activity. Aminophylline (a phosphodiesterase inhibitor) markedly elevated the retinal NAT activity, and a combined treatment with the GABA-ergic drugs and aminophylline resulted in additive effects. The actions of both muscimol and baclofen were antagonized by picrotoxin and bicuculline (two GABA-A receptor blockers), whereas the effect of baclofen was not changed by a selective GABA-B receptor blocker, CGP 35,348. Melatonin given ip significantly raised NAT activity, and its combination with muscimol further stimulated the enzyme. Picrotoxin and bicuculline given to chickens during the dark phase of 12 h light--12 h dark illumination cycle significantly suppressed the nocturnal NAT activity in retina. Neither GABA nor muscimol and baclofen significantly affected basal and forskolin (1 microM)-stimulated adenylate cyclase activity in vitro in light-exposed chicken retina. It is concluded that a GABA signal (acting through type A of GABA receptors) plays an important role in a complex mechanism regulating the rhythmic melatonin biosynthesis in vertebrate retina.

Behavioral and pharmacological effects of centrally administered aminooxyacetic acid in rats.

In the present experiment unilateral intrastriatal injections of aminooxyacetic acid (1, 2.5, 5 mumol) to freely moving animals and pentobarbital-anesthetized rats produced contralateral jerks and dose-dependent mortality, but no barrel rotation. At 10-12 days there were no significant differences in exploratory activity, passive avoidance behavior, and elevated plus-maze test in aminooxyacetic acid-treated animals as compared with controls. However freely moving animals microinjected with aminooxyacetic acid (but not the pentobarbital-pretreated group) had impaired learning activity in an active avoidance conditioning test, and showed reduced striatal concentrations of substance P and GABA. Intrastriatal injections of aminooxyacetic acid therefore result in both acute and chronic behavioral changes which are attenuated by pentobarbital anesthesia.