Dopaminergic Regulation of Cone Retinomotor Movement in Isolated Teleost Retinas: II. Modulation by γ-Aminobutyric Acid and Serotonin

In the accompanying paper we reported that 3,4-dihydroxyphenylethylamine (dopamine) induced light-adaptive retinomotor movements in teleost photoreceptors and that this effect was mediated by D2 dopamine receptors located on the photoreceptors themselves. In this study, we investigated the effects on cone retinomotor movement of three agents that have been reported by others to modulate retinal dopamine release: gamma-aminobutyric acid (GABA), 5-hydroxytryptamine (5-HT, serotonin), and melatonin. We report here that the GABA antagonists bicuculline and picrotoxin induced light-adaptive cone contraction in dark-adapted green sunfish retinas cultured in constant darkness; thus they mimic the effect of light or exogenously applied dopamine. Since their effects were blocked by either the D2 dopamine antagonist sulpiride or by Co2+, it seems likely that these agents act by enhancing retinal dopamine release. The GABA agonist muscimol produced effects opposite to those of GABA antagonists. Muscimol inhibited light-induced cone contraction in previously dark-adapted retinas and induced dark-adaptive cone elongation in light-adapted retinas. These results suggest that in green sunfish retinas, as has been reported for other retinas, GABA inhibits dopamine release. 5-HT induced light-adaptive cone contraction in dark-adapted retinas; thus 5-HT also mimics the effect of light or exogenously applied dopamine. The effect of 5-HT was blocked by sulpiride, Co2+, or the 5-HT antagonist mianserin. These results suggest that 5-HT induces cone contraction by stimulating dopamine release. Melatonin neither inhibited dopamine-induced cone contraction in retinas cultured in the dark nor induced cone elongation in retinas cultured in the light. Our results suggest that both GABA and 5-HT (but not melatonin) affect cone retinomotor movements in green sunfish by modulating dopamine release: GABA by inhibiting and 5-HT by stimulating dopamine release. We report in the companion paper that dopamine induced contraction in isolated cone fragments. Together these observations strongly suggest that dopamine serves as the final extracellular messenger directly inducing light-adaptive cone retinomotor movement, and that GABA and 5-HT affect these movements by modulating dopamine release.     

Dopaminergic Regulation of Cone Retinomotor Movement in Isolated Teleost Retinas: I. Induction of Cone Contraction Is Mediated by D2 Receptors

In the retinas of lower vertebrates, retinal photoreceptors and melanin pigment granules of the retinal pigment epithelium (RPE) undergo characteristic movements in response to changes in light intensity and to signals from an endogenous circadian clock. To identify agents responsible for mediating light and/or circadian regulation of these retinomotor movements, we investigated the effects of hormones and neurotransmitters on cone, rod, and RPE movements in the green sunfish, Lepomis cyanellus. We report here that 3,4-dihydroxyphenylethylamine (dopamine) mimics the effect of light by inducing light-adaptive retinomotor movements in all three cell types. In isolated dark-cultured retinas, dopamine induced light-adaptive cone contraction with a half-maximal effect at 10(-8) M. This effect of dopamine was inhibited by antagonists with a potency order characteristic of D2 receptor mediation. The dopamine uptake blocker benztropine also induced light-adaptive cone contraction in isolated dark-cultured retinas, suggesting that there is continuous dopamine release in the dark but that concomitant uptake normally prevents activation of cone contraction. That dopamine plays a role in light regulation of cone movement is further suggested by the observation that light-induced cone contraction was partially inhibited by sulpiride, a selective D2 dopamine antagonist, or by Co2+, a blocker of synaptic transmission. Sulpiride also promoted dark-adaptive cone elongation in isolated light-adapted retinas, suggesting that continuous dopamine action is required in the light to maintain the light-adapted cone position. Dopamine can act directly on D2 receptors located on rod and cone inner/outer segments: dopamine induced light-adaptive retinomotor movements in isolated distal fragments of dark-adapted photoreceptors cultured in the dark. Together our results indicate that dopamine induces light-adaptive retinomotor movements in cones, rods, and RPE cells by activating D2 receptors. We suggest that, in vivo, dopamine plays a role in both light and circadian regulation of retinomotor movements.     

Evidence for D4 Receptor Regulation of Retinomotor Movement in Isolated Teleost Cone Inner-Outer Segments

Abstract: In the retinas of teleost fish, cone photoreceptors change shape in response to light and circadian signals. They elongate in the dark, contract in the light, and under conditions of constant darkness undergo appropriate movements at expected dusk and dawn. Dopamine induces cones to contract, thus mimicking the effect of light or expected dawn. To identify the receptor subtype responsible for mediating dopamine regulation of cone retinomotor movements, we have carried out pharmacological studies using isolated fragments of teleost cones consisting of cone inner segments-cone outer segments (CIS-COS). Isolated CIS-COS retain the ability to elongate in dark culture and contract when subsequently exposed to light or dopamine. We report that dark-induced elongation of CIS-COS was inhibited by dopamine and its agonists with an effectiveness ranking of dopamine = quinpirole > bromocriptine ⋙SKF-38393. After 60 min of elongation in dark culture, CIS-COS myoids contracted when subsequently cultured in the dark with dopamine or quinpirole. Quinpirole-induced inhibition of elongation and quinpirole-induced contraction were completely blocked by clozapine at 1 µ M or by sulpiride at 100 µ M . These effectiveness profiles for dopamine agonists and antagonists suggest that dopamine regulation of cone retinomotor movement is mediated by a D₄-like receptor.     

Circadian regulation of teleost retinal cone movements in vitro

In the retinas of many species of lower vertebrates, retinal photoreceptors and pigment epithelium pigment granules undergo daily movements in response to both diurnal, and in the case of teleost cone photoreceptors, endogenous circadian signals. Typically, these cone movements take place at dawn and at dusk when teleosts are maintained on a cyclic light (LD) regime, and at expected dawn and expected dusk when animals are maintained in continuous darkness (DD). Because these movements are so strictly controlled, they provide an overt indicator of the stage of the underlying clock mechanism. In this study we report that both light-induced and circadian-driven cone myoid movements in the Midas cichlid (Cichlasoma citrinellum), occur normally in vitro. Many of the features of retinomotor movements found in vivo also occur in our culture conditions, including responses to light and circadian stimuli and dopamine. Circadian induced predawn contraction and maintenance of expected day position in response to circadian modulation, are also normal. Our studies suggest that circadian regulation of cone myoid movement in vitro is mediated locally by dopamine, acting via a D2 receptor. Cone myoid contraction can be induced at midnight and expected mid-day by dark culture with dopamine or the D2 receptor agonist LY171555. Further, circadian induced predawn contraction can be increased with either dopamine or LY171555, or may be reversed with the dopamine D2 antagonist, sulpiride. Sulpiride will also induce cone myoid elongation in retinal cultures at expected mid- day, but will not induce cone myoid elongation at dusk. In contrast, circadian cone myoid movements in vitro were unaffected by the D1 receptor agonist SCH23390, or the D1 receptor antagonist SKF38393. Our short-term culture experiments indicate that circadian regulation of immediate cone myoid movement does not require humoral control but is regulated locally within the retina. The inclusion of dopamine, or dopamine receptor agonists and antagonists in our cultures, has indicated that retinal circadian regulation may be mediated by endogenously produced dopamine, which acts via a D2 mechanism.     

Dopamine Induces Light-Adaptive Retinomotor Movements in Bullfrog Cones via D2 Receptors and in Retinal Pigment Epithelium via D1 Receptors

In the eyes of lower vertebrates, retinal photoreceptors and melanin pigment granules of the retinal pigment epithelium (RPE) exhibit characteristic retinomotor movements in response to changes in ambient illumination and to signals from an endogenous circadian clock. We previously reported that 3,4-dihydroxyphenylethylamine (dopamine) mimicked the effect of light on these movements in photo-receptors and RPE cells of green sunfish, Lepomis cyanellus, by interacting with D2 dopaminergic receptors. Here, we report that dopamine also mimics the effect of light on cone and RPE retinomotor movements in bullfrogs, Rana catesbeiana, i.e., dopamine induces cone contraction and RPE pigment dispersion. Dopamine induced cone contraction in isolated dark-adapted bullfrog retinas incubated in constant darkness in the presence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX). This effect of dopamine was inhibited by a D2 but not a D1 antagonist and mimicked by a D2 but not a D1 agonist. These results suggest that induction of cone contraction by dopamine is mediated by D2 dopaminergic receptors and that cone adenylate cyclase activity is inhibited. Thus, dopamine acts via the same type of receptor in both bullfrog and green sunfish retinas to induce cone contraction. In contrast, dopamine influences RPE retinomotor movement via different receptors in fish and bullfrog. Dopamine induced light-adaptive pigment dispersion in isolated dark-adapted bullfrog RPE-eyecups incubated in constant darkness in normal Ringer's solution. Because the retina was not present, these experiments demonstrate a direct effect of dopamine on bullfrog RPE. This effect of dopamine on bullfrog RPE was inhibited by a D1 but not a D2 antagonist and mimicked by a D1 but not a D2 agonist. Furthermore, agents that increase the concentration of intracellular cyclic AMP also induced pigment dispersion in dark-adapted bullfrog RPE-eyecups incubated in the dark. These results suggest that dopamine induces pigment dispersion in bullfrog RPE via D1 dopaminergic receptors. Thus, dopamine acts via different receptors on bullfrog (D1) versus green sunfish (D2) RPE to induce pigment dispersion. In addition, inhibitor studies indicate that pigment dispersion is actin dependent in teleost but not in bullfrog RPE. Dopamine-induced pigment dispersion was inhibited by cytochalasin D in isolated RPE sheets of green sunfish but not in RPE-eyecups of bullfrogs. Together, these observations indicate that dopamine mimics the effect of light on cone and RPE retinomotor movements in both fish and bullfrogs. However, in the RPE, different receptors mediate the effect of dopamine, and different cytoskeletal mechanisms are used to affect pigment transport.(ABSTRACT TRUNCATED AT 400 WORDS)     

Adenosine Stimulates Cone Photoreceptor Myoid Elongation via an Adenosine A2-Like Receptor

In several parts of the nervous system, adenosine has been shown to function as an extracellular neuromodulator binding to surface receptors on target cells. This study examines the possible role of adenosine in mediating light and circadian regulation of retinomotor movements in teleost cone photoreceptors. Teleost cones elongate in the dark and contract in the light. In continuous darkness, the cones continue to elongate and contract at subjective dusk and dawn in response to circadian signals. We report here that exogenous adenosine triggers elongation (the dark/night movement) in isolated cone inner segment-cone outer segment preparations (CIS-COS) in vitro. Agonist/antagonist potency profiles indicate that adenosine's effect on cone movement is mediated by an A2-like adenosine receptor, which like other A2 receptors enhances adenylate cyclase activity. Although closest to that expected for A2 receptors, the antagonist potency profile for CIS-COS does not correspond exactly to any known A2 receptor subtype, suggesting that the cone receptor may be a novel A2 subtype. Our findings are consistent with previous reports that retinal adenosine levels are higher in the dark, and further suggest that adenosine could act as a neuromodulatory "dark signal" influencing photoreceptor metabolism and function in the fish retina.     

Taurine blocks spontaneous cone contraction but not horizontal cell dark suppression in isolated goldfish retina

The objective of this study was to investigate the effects of taurine on cone retinomotor movements and the responses of cone-driven horizontal cells in dark-adapted teleost retina. In isolated goldfish retina preparations maintained in the dark, cones spontaneously contracted, and the responses of horizontal cells were suppressed. Addition of 5 mM taurine to the physiological solution blocked the spontaneous contraction of cones in the dark but did not block the dark-suppression of horizontal cell responses. These results indicate that the mechanism that leads to horizontal cell dark suppression is not sensitive to taurine. Although both cone retinomotor position and horizontal cell responsiveness are known to be modulated by dopamine, the present results do not support the hypothesis that taurine inhibits dopamine release in the dark because only spontaneous cone contraction was affected by taurine. These results also indicate that spontaneous cone contraction in the dark is not the cause of horizontal cell dark suppression because, in the presence of taurine, cones were elongated yet horizontal cell responses were still suppressed. Consequently, these results make it clear that horizontal cell dark suppression is not an artifact produced by incubating isolated teleost retina preparations in taurine-free physiological solution.     

Effects of cyclic adenosine 3'',5''-monophosphate on photoreceptor disc shedding and retinomotor movement. Inhibition of rod shedding and stimulation of cone elongation

As a test of the hypothesis that cyclic nucleotides play a role in the regulation of retinomotor movements and disc shedding in the photoreceptor-pigment epithelial complex, we have used an in vitro eyecup preparation that sustains both disc shedding and cone retinomotor movements, Eyecups were prepared in white light from animals in which both shedding and cone movement had been blocked by 4 d of constant-light treatment. In eyecups incubated for 3 h in light, disc shedding was negligible and cones remained in the light-adapted (contracted) position. In eyecups incubated in darkness, however, a massive shedding response (dominated by rod photoreceptors) was induced, and at the same time cone photoreceptors elongated to their dark-adapted position. In eyecups incubated in light dbcAMP promoted cone elongation and thus mimicked darkness; the dbcAMP effect was potentiated by the phosphodiesterase inhibitors papaverine and 3- isobutylmethylxanthine. In eyecups incubated in darkness, on the other hand, both phosphodiesterase inhibitors and dbcAMP reduced the phagosome content of the pigment epithelium. The effects of dbcAMP on the cone elongation and rod shedding appear to be specific in that dbcGMP, adenosine, and adenosine 5'-monophosphate had no significant effect. Our results suggest that cAMP plays a role in the regulation of both retinomotor movements and disc shedding.     

Stimulation of Distinct D2 Dopaminergic and α2-Adrenergic Receptors Induces Light-Adaptive Pigment Dispersion in Teleost Retinal Pigment Epithelium

In the retinal pigment epithelium (RPE) of lower vertebrates, melanin pigment granules aggregate and disperse in response to changes in light conditions. Pigment granules aggregate into the RPE cell body in the dark and disperse into the long apical projections in the light. Pigment granule movement retains its light sensitivity in vitro only if RPE is explanted together with neural retina. In the absence of retina, RPE pigment granules no longer move in response to light onset or offset. Using a preparation of mechanically isolated fragments of RPE from green sunfish, Lepomis cyanellus, we investigated the effects of catecholamines on pigment migration. We report here that 3,4-dihydoxyphenylethylamine (dopamine) and clonidine each mimic the effect of light in vivo by inducing pigment granule dispersion. Dopamine had a half-maximal effect at approximately 2 nM; clonidine, at 1 microM. Dopamine-induced dispersion was inhibited by the D2 dopaminergic antagonist sulpiride but not by D1 or alpha-adrenergic antagonists. Furthermore, a D2 dopaminergic agonist (LY 171555) but not a D1 dopaminergic agonist (SKF 38393) mimicked the effect of dopamine. Clonidine-induced dispersion was inhibited by the alpha 2-adrenergic antagonist yohimbine but not by sulpiride. These results suggest that teleost RPE cells possess distinct D2 dopaminergic and alpha 2-adrenergic receptors, and that stimulation of either receptor type is sufficient to induce pigment granule dispersion. In addition, forskolin, an activator of adenylate cyclase, induced pigment granule movement in the opposite direction, i.e., dark-adaptive pigment aggregation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of extracellular Ca++, K+, and Na+ on cone and retinal pigment epithelium retinomotor movements in isolated teleost retinas

We have examined the effects of changes in extracellular ionic composition on cone and retinal pigment epithelium (RPE) retinomotor movements in cultured isolated teleost retinas. In vivo, the myoid portion of teleost cones contracts in the light and elongates in the dark; RPE pigment disperses in the light and aggregates in the dark. In vitro, cones of dark-adapted (DA) retinas cultured in constant darkness contracted spontaneously to their light-adapted (LA) positions if the culture medium contained greater than or equal to 10(-3)M Cao++. DA cones retained their long DA positions in a medium containing less than or equal to 10(-6)M Cao++. Low [Ca++]o (10(-5)-10(-7)M) also permitted darkness to induce cone elongation and RPE pigment aggregation. Light produced cone contraction even in the absence of Cao++, but the extent of contraction was reduced if [Ca++]o was less than 10(-3) M. Thus, full contraction appeared to require the presence of external Ca++. High [K+]o (greater than or equal to 27 mM) inhibited both light-induced and light-independent Ca++-induced cone contraction. However, low [Na+]o (3.5 mM) in the presence of less than or equal to 10(-6)M Cao++ did not mimic light onset by inducing cone contraction in the dark. High [K+]o also promoted dark-adaptive cone and RPE movements in LA retinas cultured in the light. All results obtained in high [K+]o were similar to those observed when DA or LA retinas were exposed to treatments that elevate cytoplasmic cyclic 3',5'-adenosine monophosphate (cAMP) content.     

Dopamine Inhibits Forskolin- and 3-Isobutyl-1-Methylxanthine-Induced Dark-Adaptive Retinomotor Movements in Isolated Teleost Retinas

We have been investigating the mechanisms of diurnal and circadian regulation of teleost retinomotor movements. In the retinas of lower vertebrates, photoreceptors and melanin pigment granules of the retinal pigment epithelium (RPE) undergo movements at dawn and dusk. These movements continue to occur at subjective dawn and dusk in animals maintained in constant darkness. Cone myoids contract at dawn and elongate at dusk; RPE pigment disperses into the epithelial cells' long apical processes at dawn and aggregates into the cell bodies at dusk. We report here that forskolin, an adenylate cyclase activator, and 3-isobutyl-1-methylxanthine (IBMX), a phosphodiesterase inhibitor, each induces dark-adaptive cone and RPE retinomotor movements in isolated light-adapted green sunfish retinas cultured in constant light. Forskolin induces a 22-fold elevation in retinal cyclic AMP content. Forskolin- and IBMX-induced movements are inhibited approximately 65% and 95%, respectively, by 3,4-dihydroxyphenylethylamine (dopamine). However, dopamine does not inhibit dark-adaptive movements induced by dibutyryl cyclic AMP. Epinephrine is much less effective than dopamine in inhibiting forskolin-induced movements, while phenylephrine and clonidine are totally ineffective. These results are consistent with our previous findings that treatments that increase intracellular cyclic AMP content promote dark-adaptive retinomotor movement. They further suggest that dopamine inhibits adenylate cyclase activity in photoreceptors and RPE cells and thereby favors light-adaptive retinomotor movements.     

Differential G protein-mediated coupling of D2 dopamine receptors to K+ and Ca2+ currents in rat anterior pituitary cells.

In anterior pituitary cells, dopamine, acting on D2 dopamine receptors, concomitantly reduces calcium currents and increases potassium currents. These dopamine effects require the presence of intracellular GTP and are blocked by pretreatment of the cells with pertussis toxin, suggesting that one or more G protein is involved. To identify the G proteins involved in coupling D2 receptors to these currents, we performed patch-clamp recordings in the whole-cell configuration using pipettes containing affinity-purified polyclonal antibodies raised against either Go alpha, Gi3 alpha, or Gi1,2 alpha. Dialysis with Go alpha antiserum significantly reduced the inhibition of calcium currents induced by dopamine, while increase of potassium currents was markedly attenuated only by Gi3 alpha antiserum. We therefore conclude that in pituitary cells, two different G proteins are involved in the signal transduction mechanism that links D2 receptor activation to a specific modulation of the four types of ionic channels studied here.     

An islet activating protein-sensitive G protein is involved in dopamine inhibition of angiotensin and thyrotropin-releasing hormone-stimulated inositol phosphate production in anterior pituitary cells

In primary culture of anterior pituitary cells, dopamine inhibited the angiotensin (AII)-stimulated inositol phosphate production by 28 +/- 2.5% (n = 14), with an EC50 of 660 +/- 228 nM (n = 8). This effect was blocked by (+)-butaclamol, a specific dopamine receptor antagonist. RU 24926, a D2 specific agonist, but not SKF 38393, a specific D1 agonist, inhibited AII-stimulated inositol phosphate production, suggesting that this dopamine effect is mediated through a dopamine receptor of the D2 subtype. Dopamine also partially inhibited (25%) inositol phosphate production stimulated by thyrotropin-releasing hormone (TRH). Our results suggest that the dopamine-mediated inhibition of hormonally stimulated inositol phosphate production is probably not mediated through the known inhibitory effects of dopamine on cAMP and Ca2+ intracellular concentrations. Although unknown, the mechanism by which dopamine inhibited the AII and TRH-stimulated inositol phosphate production implicates a GTP binding protein sensitive to the islet activating protein (IAP) since dopamine effects were blocked by this toxin. The alpha subunit of the GTP binding protein involved could be one of the three ADP-ribosylated proteins found in anterior pituitary cells in primary cultures, the alpha o (39 kDa), the alpha i (41 kDa), and an alpha subunit of 40 kDa. Indeed, we show here that this 40-kDa IAP substrate, already described in a few tissues, is present in anterior pituitary cells. The negative coupling between dopamine receptors and the AII or TRH inositol phosphate production systems, could be implicated in the dopamine inhibition of the AII- and TRH-stimulated prolactin release since such an inhibition is blocked by IAP. Our results suggest that the negative regulation of inositol phosphate production is one of the mechanisms by which dopamine controls hormonally stimulated prolactin release.     

Evidence for a D2 dopamine receptor in frog retina that decreases cyclic AMP accumulation and serotonin N-acetyltransferase activity

The regulation of serotonin N-acetyltransferase (NAT) activity and cyclic AMP accumulation in the retina of the African clawed frog (Xenopus laevis) was studied using an in vitro eye cup preparation. Retinal NAT, a key enzyme in the synthesis of melatonin, is expressed as a circadian rhythm with peak activity at night. The increase of NAT activity at night appears to be mediated by cyclic AMP and is suppressed by light. Dopamine inhibits the nocturnal increase of retinal NAT activity; approximately 80% inhibition was observed with 1 microM dopamine. Dopamine at 1 microM did not stimulate retinal cyclic AMP accumulation. The effect of dopamine on NAT activity was antagonized by the D2-selective receptor antagonists spiperone and metoclopramide, but not by the putative D1 selective antagonist SCH 23390. The nocturnal rise in NAT activity was inhibited by LY 171555, a putative D2 selective agonist, but not by SKF 38393, a putative D1 selective agonist. LY 171555 also decreased cyclic AMP accumulation in eye cups incubated under similar conditions. Dopamine inhibited the stimulation of NAT activity in light by 3-isobutylmethylxanthine, but not that by dibutyryl cyclic AMP, suggesting that dopamine acts by decreasing cyclic AMP formation in the NAT-containing cells. Thus, the effects of dopamine on NAT activity may be mediated by a receptor with the pharmacological and biochemical characteristics of a D2 receptor.     

D2-dopamine receptor and alpha2-adrenoreceptor-mediated analgesic response of quercetin

Quercetin, a bioflavonoid (100-300 mg/kg) produced dose dependent increase in tail-flick latency, the analgesic effect being sensitive to reversal by naloxone (1 mg/kg). Prior treatment with haloperidol (1 mg/kg), D1/D2 receptor antagonist haloperidol, sulpiride (50 mg/kg), a selective D2 receptor antagonist, yohimbine (5 mg/kg), a alpha2-adrenoreceptor antagonist but not by SCH 23390 a, selective D1 receptor antagonist blocked this response. Apomorphine (1 mg/kg) a mixed D1/D2 dopamine receptor agonist, and quinpirole (0.5 mg/kg), a selective D2 receptor agonist also produced antinociception, that was reversed by haloperidol (1 mg/kg), sulpiride (50 mg/kg), but not by yohimbine (5 mg/kg). The antinociceptive action of quercetin (200 mg/kg) was potentiated by D2 agonist quinpirole (0.2 mg/kg). Dopamine D1 receptor agonist SKF38393 (10 and 15 mg/kg) failed to alter the antinociceptive effect of quercetin (200 mg/kg). Quercetin (200 mg/kg) reversed reserpine (2 mg/kg-4 hr) induced hyperalgesia, which was reversed by sulpiride but not by yohimbine. Thus, a role of dopamine D2 and alpha2-adrenoreceptors is postulated in the antinociceptive action of quercetin.     

Aromatic L-Amino Acid Decarboxylase Is Modulated by D1 Dopamine Receptors in Rat Retina

Aromatic L-amino acid decarboxylase (AAAD) activity of rat retina increases when animals are placed in a lighted environment from the dark. The increase of activity can be inhibited by administering the selective dopamine D1 receptor agonist SKF 38393, but not the selective D2 agonist quinpirole, or apomorphine. Conversely, in the dark, enzyme activity can be enhanced by administering the selective D1 antagonist SCH 23390 or haloperidol, but not the selective D2 antagonist (-)-sulpiride. Furthermore, in animals exposed to room light for 3 h, the D1 agonist SKF 38393 reduced retinal AAAD activity, and this effect was prevented by prior administration of SCH 23390. In contrast, quinpirole had little or no effect when administered to animals in the light. Kinetic analysis indicated that the apparent Vmax for the enzyme increases with little change in the apparent Km for the substrate 3,4-dihydroxyphenylalanine or the cofactor pyridoxal-5'-phosphate. We suggest that dopamine released in the dark tonically occupies D1 receptors and suppresses AAAD activity. When the room light is turned on, D1 receptors are vacated and selective D1 agonists can either prevent the rise of AAAD or reverse light-enhanced AAAD activity.     

Dopamine inhibits mitochondrial motility in hippocampal neurons

Background

The trafficking of mitochondria within neurons is a highly regulated process. In an earlier study, we found that serotonin (5-HT), acting through the 5-HT1A receptor subtype, promotes axonal transport of mitochondria in cultured hippocampal neurons by increasing Akt activity, and consequently decreasing glycogen synthase kinase (GSK3β) activity. This finding suggests a critical role for neuromodulators in the regulation of mitochondrial trafficking in neurons. In the present study, we investigate the effects of a second important neuromodulator, dopamine, on mitochondrial transport in hippocampal neurons.

Methodology/Principal Findings

Here, we show that dopamine, like 5-HT, regulates mitochondrial motility in cultured hippocampal neurons through the Akt-GSK3β signaling cascade. But, in contrast to the stimulatory effect of 5-HT, administration of exogenous dopamine or bromocriptine, a dopamine 2 receptor (D2R) agonist, caused an inhibition of mitochondrial movement. Moreover, pretreatment with bromocriptine blocked the stimulatory effect of 5-HT on mitochondrial movement. Conversely, in cells pretreated with 5-HT, no further increases in movement were observed after administration of haloperidol, a D2R antagonist. In contrast to the effect of the D2R agonist, addition of {"type":"entrez-protein","attrs":{"text":"SKF38393","term_id":"1157151916","term_text":"SKF38393"}}SKF38393, a dopamine 1 receptor (D1R) agonist, promoted mitochondrial transport, indicating that the inhibitory effect of dopamine was actually the net summation of opposing influences of the two receptor subtypes. The most pronounced effect of dopamine signals was on mitochondria that were already moving directionally. Western blot analysis revealed that treatment with either a D2R agonist or a D1R antagonist decreased Akt activity, and conversely, treatment with either a D2R antagonist or a D1R agonist increased Akt activity.

Conclusions/Significance

Our observations strongly suggest a role for both dopamine and 5-HT in regulating mitochondrial movement, and indicate that the integrated effects of these two neuromodulators may be important in determining the distribution of energy sources in neurons.     

Effect of melatonin agonists and antagonists on horizontal cell spinule formation and dopamine release in a fish retina

The crucian carp retina was used to study the effects of the melatonin antagonist p697 (N-pentanoyl 2-benzyltryptamine) and the melatonin agonists [+]- and [-]-AMMTC (N-acetyl-4-aminomethyl-6-methoxy-9-methyl-1,2,3,4-tetrahydrocarbazol e) on horizontal cell spinule formation, an indicator of the state of retinal adaptation. DH97 was capable of both counteracting dark-adaptive spinule degradation and inducing light-adaptive spinule formation at the beginning of the dark phase. Addition of dopamine receptor blockers opposed the action of DH97 on spinules, with SCH 23930, a D1 dopamine receptor antagonist, being more effective than the D2 receptor antagonist sulpiride. DH97 induced a twofold increase in dopamine release. We conclude that melatonin acts as a dark signal within the teleost retina by inhibiting the dopaminergic system. In accordance with this, both enantiomers of AMMTC prevented light-induced spinule formation, and reduced dopamine release to below dark-adaptive baseline levels. We suggest that the suppression of spinule formation by AMMTC may be due to either a direct inhibitory interaction between the melatonin agonist and horizontal cell dopamine D1 receptors, or an inhibitory effect on the activity of the dopamine-releasing interplexiform cells.