Calcium channel blockers in vivo inhibit serotonin N-acetyltransferase (NAT) activity in chicken retina stimulated by darkness and not by agents elevating intracellular cyclic AMP level.

The molecular mechanism underlying the role of calcium influx in the regulation of retinal serotonin N-acetyltransferase (NAT) activity was studied in vivo in chickens. Systemic administration of organic antagonists of voltage-sensitive calcium channels (VSCC), i.e., nimodipine and nifedipine, resulted in a marked suppression of the nocturnal increase of NAT activity in chicken retina. In contrast, NAT activity stimulated by nonhydrolysable analogs of cyclic AMP (dibutyryl-cyclic AMP and 8-bromo-cyclic AMP), forskolin, a direct activator of adenylate cyclase, and by phosphodiesterase inhibitors (aminophylline and 3-isobutyl-1-methylxanthine), was not significantly affected by various tested VSCC antagonists. The inhibitory effect of nimodipine on the dark-dependent increase in NAT activity of chicken retina was abolished by Bay K 8644, a selective VSCC agonist. The results presented in this paper indicate an important role of calcium influx through L-type of VSCC in the induction of NAT activity in chicken retina, and suggest that a requirement of calcium ions in the process of NAT induction in the retina may be primarily at the level of cyclic AMP production.

Serotonin N-acetyltransferase (NAT) induction in mammalian retina: role of cyclic AMP and calcium ions.

In retinas and pineal glands of rat, rabbit and hen, activities of the penultimate (and key regulatory) enzyme in melatonin biosynthesis, serotonin N-acetyltransferase (NAT), display distinct diurnal variations, with high and low values during dark and light phase of a 12-h dark: 12-h light illumination cycle. Two-hour incubation (during daytime hours in light) of isolated pineal glands of the studied vertebrates, or the retinas, with 50 microM forskolin (plus 100 microM 3-isobutyl-1-methylxanthine, IBMX-a phosphodiesterase inhibitor), and 1 mM dibutyryl-cAMP, markedly increased the tissue NAT activity. The same procedures significantly enhanced the enzyme activity of rat retina in light, however, only during nighttime hours. The forskolin (+ IBMX)-induced increase of NAT activity in rat retina was significantly lower in a calcium-free medium, and substantially enhanced when calcium concentration was raised from 1.3 mM to 3.9 mM. Treatment of rats with IBMX or aminophylline, and rabbits with aminophylline, increased NAT activity in their pineal glands irrespective of the time of the day, whereas both phosphodiesterase inhibitors significantly increased the enzyme activity of rat retina only when injected during the subjective dark hours. It is concluded that, by analogy to vertebrate pineal gland, in vertebrate retina an increase of NAT activity (and consequently melatonin formation), stimulated both physiologically (i. e. at night), or pharmacologically, involves a cAMP- and calcium dependent process of the enzyme induction.

Serotonin N-acetyltransferase activity in chicken retina: in vivo effects of phosphodiesterase inhibitors, forskolin, and drugs affecting dopamine receptors.

A role of D2-dopaminergic neurotransmission in the regulation of melatonin biosynthesis in retina was studied in vivo in chickens. The nighttime rise in serotonin N-acetyltransferase (NAT)--the penultimate and key regulatory melatonin-synthesizing enzyme--was potently inhibited by both acute light exposure and agonists of dopamine D2-receptor (quinpirole, bromocriptine, and apomorphine). Spiroperidol, a selective dopamine D2-receptor blocker, increased the enzyme activity in light-exposed chickens, but had no effect in animals kept in darkness. Inhibitors of cyclic nucleotide phosphodiesterase, aminophylline, and 3-isobutyl-1-methylxanthine given peripherally, along with a direct adenylate cyclase activator forskolin injected directly into the eye, mimicked the action of darkness, and markedly enhanced the retinal NAT activity when administered to animals maintained in an illuminated environment. Dopamine D2-receptor agonists had no effect on aminophylline-stimulated enzyme activity, whereas spiroperidol enhanced it. Forskolin-driven NAT activity was suppressed by quinpirole. Spiroperidol and aminophylline given alone at different times of day under light conditions stimulated NAT activity, and their effects were mainly additive when given in combination. SCH 23390, a selective D1-dopamine receptor antagonist, did not affect the rise in NAT activity of chicken retina produced by either darkness or by aminophylline. The results provide further evidence that dopamine, acting via D2-receptors, mediates the inhibitory effects of light on the cyclic AMP-dependent dark-evoked induction of NAT activity in chicken retina.

Activation of D2 dopamine receptors in hen retina decreases forskolin-stimulated cyclic AMP accumulation and serotonin N-acetyltransferase (NAT) activity.

Dopamine (DA; 1-100 ?M) alone did not significantly stimulate cyclic AMP accumulation in the hen retina; however, in the presence of 0.3 ?M spiroperidol, it clearly increased it. DA, in a concentration-dependent manner, enhanced the stimulatory effect of forskolin (1 and 10 ?M), both in the absence and presence of a phosphodiesterase (PDE) inhibitor IBMX (0.5 mM). A selective D1-receptor blocker such as SCH 23390 antagonized the activating effect of DA when used at 1 ?M, and reversed the amine action from a stimulatory into an inhibitory effect when applied at 3 or 10 ?M concentration. Addition of 0.3 ?M spiroperidol further enhanced the DA action on cyclic AMP accumulation evoked by forskolin; however, spiroperidol used at 5 ?M concentration antagonized the potentiating effect of the amine. Selective agonists of DA D2-receptors such as quinpirole (LY 1715550; 0.01-10 ?M) and bromocriptine (1-100 ?M) decreased both basal levels and forskolin-stimulated cyclic AMP accumulation in a concentration-dependent manner in the absence and presence of 0.5 mM IBMX. Under daylight conditions, dibutyryl-cyclic AMP (1 mM) and a combination of 50?M forskolin and 0.1 mM IBMX (these drugs applied separately were only weakly active) significantly enhanced the activity of serotonin N-acetyltransferase (NAT) activity (the key regulatory enzyme in melatonin biosynthesis) in pieces of the hen retina. Quinpirole (0.001-10 ?M) inhibited the effect of forskolin (combined with IBMX) in a concentration-dependent and spiroperidol-sensitive manner, and did not modify the action of dibutyrylcyclic AMP. It is concluded that the hen retina possesses both types of DA receptors, i.e. D1 and D2, whose stimulation respectively increase and decrease cyclic AMP levels. These D2-receptors, which are negatively coupled to adenylate cyclase, seem to be involved in the regulation of NAT activity in the retina, and their activation leads to the inhibition of the enzyme induction.

Dopamine in the rabbit retina and striatum: diurnal rhythm and effect of light stimulation.

In rabbits, dopamine levels in the retina, but not in the caudate nucleus, showed clear diurnal rhythm, with high values seen in the light phase. Thirty min exposition of dark-adapted rabbits to day-light produced no changes in dopamine levels in the retina. In rabbits treated with alpha-methyl-p-tyrosine, the same light exposition decreased the retinal amine level by 18%, while stimulation with intensive, flickering light significantly decreased the retinal dopamine content by 36%. Experiments performed at noon and midnight, under light or dark conditions, showed the retinal dopamine levels to be very similar in groups kept either at light or dark, irrespective of the time of the day, although in animals deprived of light the amine levels were clearly lower than in those exposed to light, both at noon and midnight. Under all experimental conditions there were no significant changes in dopamine level and utilization in the caudate nucleus. The isolated and superfused retina (preloaded with [3H]-dopamine), when stimulated with flashes of white light (2 Hz, 10 min), released [3H]-radioactivity in a Ca2+-dependent manner. It is concluded that in rabbits, light enhances dopamine levels and utilization selectively in the retina, and the observed diurnal changes in the amine metabolism are dependent on the presence or absence of light, and not on the time of the day. The proposed physiological role(s) of the retinal dopaminergic mechanisms is discussed.

Serotonin N-acetyltransferase (NAT) activity in hen retina and pineal gland: in vivo pharmacological induction at noon and antagonism of the light-evoked suppression at night.

In vivo effective pharmacological procedures are described which markedly increase activity of serotonin N-acetyltransferase (NAT), the key regulatory enzyme in melatonin biosynthesis, during the daytime (in light) and counteract suppressive effects of light on NAI activity at night in the hen retina and pineal gland. Of the tested compounds, and their combinations, the most effective were: "aminophylline + spiroperidol + alpha-methyl-p-tyrosine" for the retina, and "aminophylline + yohimbine (+ alpha-methyl-p-tyrosine)" for the pineal gland. The results give strong support to the concept that the dopaminergic (C(2)-receptor) and noradrenergic (alpha(2)-adrenergic receptor) mechanisms control NAT activity, and melatonin synthesis, in the hen retina and pineal gland, respectively.

The melatonin generating system in the rat retina and pineal gland: effect of single and repeated electroconvulsive shock (ECS).

N-Acetyltransferase (NAT), a rate-limiting enzyme in the melatonin synthesis which converts serotonin to N-acetylserotonin, shows a distinct circadian rhythm in the rat pineal gland and retina, with low activities during the light phase and peak activities during the dark phase. Hydroxyindole-O-methyltransferase (HIOMT), an enzyme which methylates N-acetylserotonin to melatonin, did not show any significant diurnal variations in both analyzed tissues. Isoproterenol, a selective beta-adrenoceptor agonist, when administered during morning hours of the light phase, markedly increased NAT activity in the pineal gland, but not in the retina. Electroconvulsive shock (ECS), especially when applied repeatedly (ECS x 10, once daily) significantly increased NAT activity in the retina and tended to decrease the enzyme activity in the pineal gland in isoproterenol-treated rats. ECS x 10 slightly increased and decreased the nocturnally-stimulated NAT activity in the rat retina and pineal gland, respectively.