Precocious induction of luteal activation and termination of delayed implantation in mink with the dopamine antagonist pimozide

The effects of the dopamine antagonist pimozide on the preimplantation delay phase of mink gestation were investigated in field and laboratory trials. Three doses of 0.1 mg pimozide in acetic acid administered on the 7th, 9th and 11th days after mating abbreviated gestation in Pastel kit female mink to a mean (+/- SEM) of 45.5 +/- 0.5 days, 10 days less than that observed in mink treated with vehicle only (55.6 +/- 0.6 days). In laboratory trials, four doses of 0.1 mg pimozide on the 7th, 9th, 11th and 13th day after mating resulted in embryo implantation at a mean of 25 +/- 4.3 days after mating while vehicle-treated control animals had mean preimplantation delay of 37 +/- 3.1 days. Luteal activation in the pimozide-treated group, as indicated by a rapid increase in circulating progesterone, began within 2 days after the first pimozide injection. No increase was observed in vehicle-treated mink until 6 or more days after the initiation of injections or 13 days after mating. It was concluded that pimozide, presumably by permitting endogenous secretion of prolactin, can induce precocious luteal activation and embryo implantation in the mink.     

Tail Regeneration in Normal,Blinded and Pinealectomized Gekkonid Lizards,Hemidactylus flaviviridis,Exposed to Four Different Light Conditions During Three Seasons (Temperatures)

Tail regeneration was followed for 60 days in 1470 normal (NL), blinded (BL) and pinealectomized (PX) gekkonid lizards, Hemidactylus flaviviridis, fed on cockroaches ad libitum and exposed to continuous light of high intensity (2500 lux), continuous light of low intensity (638 lux), 12 h of light (high intensity) and 12 h of darkness and continuous darkness in summer (March – May; cage temperature 30°C) monsoon (August – October; cage temperature 26°C) and winter (November – January; cage temperature 17°C) seasons. A comparative assessment of the new growth (regenerate) showed that the initiation of regeneration, the daily growth, the total length of tail replaced at the end of regeneration and the total percentage replacement of the lost (autotomized) tails were all enhanced by continuous light and depressed by continuous darkness. Furthermore, there was no significant difference between any of the parameters in NL and BL Hemidactylus; however, PX lizards generally showed retardation in the regeneration process. Seasonal differences in tail regeneration were noted—the best regenerative performance was obtained during the summer months and the worst during the winter months, with the regenerative performance during the monsoon season in between. Thermal and photoperiodic influences on tail regeneration in lizards are discussed.     

Possible dopaminergic system involvement in LH and PRL responses to naltrexone treatment

Naltrexone (Nalt) causes a rapid increase in luteinizing hormone (LH) level. This short term increase of LH concentration declines to baseline levels in less than 1 hour. Addition of pimozide (0.1 mg) caused a blunted response to Nalt challenge, with significantly reduced LH peak values compared with Nalt treatment alone. Pimozide alone caused a delayed decrease compared with baseline LH values. By following plasma prolactin (PRL) levels it was shown that pimozide administration increased PRL levels rapidly for more than 2 hours. Addition of Nalt to pimozide-treated rats significantly decreased plasma PRL values compared with pimozide alone. Nalt injected by itself attenuated PRL baseline levels. Thus, the mechanism by which pimozide caused PRL elevated level is via the dopaminergic as well as the opioid system. It is suggested that the opioid system controls plasma PRL and LH levels through other hypothalamic neurotransmitters in addition to dopamine.     

ALTERATIONS IN RECEPTORS CONTROLLING DOPAMINE SYNTHESIS AFTER CHRONIC ETHANOL INGESTION

The influence of acute and chronic ethanol treatment and withdrawal on regulation of dopamine synthesis in striatal and mesolimbic areas of mouse brain was evaluated. Tyrosine hydroxylase activity was estimated by measuring in vivo DOPA accumulation after inhibition of aromatic amino acid decarboxylase. Eight hours after a single (3 g/kg) dose of ethanol, DOPA synthesis was increased and pimozide, a dopamine receptor antagonist, stimulated DOPA synthesis to the same degree in ethanol-treated and control animals. On the other hand, 8 h after withdrawal of animals from chronic ethanol treatment, endogenous dopamine synthesis was the same in ethanol-withdrawn and control animals, but the stimulation of dopamine synthesis produced by low doses of pimozide or haloperidol was significantly less in the animals that had consumed ethanol. This effect was even more apparent at 24h after withdrawal; by 3 days after withdrawal the decreased response of ethanol-withdrawn animals to the administration of dopamine receptor blockers was no longer statistically significant. At all time points tested, high doses of pimozide or haloperidol stimulated DOPA synthesis equally in control and ethanol-withdrawn animals. Chronic ethanol treatment and withdrawal may alter the coupling between dopamine receptors which regulate dopamine synthesis and tyrosine hydroxylase.     

Morphine can stimulate prolactin release independent of a dopaminergic mechanism

Prolactin release is controlled by prolactin-release inhibiting factor (PIF), possibly dopamine, and an unidentified putative hypothalamic prolactin-releasing factor (PRF). Morphine and related opioids may indirectly stimulate prolactin release by inhibiting PIF release and (or) by stimulating putative PRF release. In the present study, we have completely blocked the dopaminergic receptors in normal male rats by pretreatment with a large dose of pimozide (3 mg/kg) to demonstrate if putative PRF has a role in morphine-induced prolactin release. Morphine sulfate (10 mg/kg) was still able to stimulate prolactin release in the rat without any functional dopaminergic PIF receptors. When naloxone (3 mg/kg) was injected 20 min before the morphine in the pimozide-treated rat, plasma prolactin concentration was not affected by morphine indicating that the stimulatory effect of this opioid on prolactin release in the pimozide-pretreated rat was mediated by mu-receptors. We can conclude that morphine can stimulate prolactin release through a mechanism apparently independent of dopaminergic receptors, one possible route being through a putative PRF.     

Tail regeneration in the lizards Anguis fragilis and Lacerta dugesii

Rates of tail regeneration in the Madeira wall lizard ( Lacerta dugesii ) and the slow-worm ( Anguis fragilis ) were studied.
L. dugesii regenerates very rapidly, the new tail sometimes attaining a maximum rate of growth of 2'6 mm a day during the fifth week after autotomy. By the twelfth week 90% of the original tail length has been replaced. Average regeneration rates of samples of lizards were reduced after repeated autotomies, but our investigation of this problem was probably complicated by another factor, the amount of tail lost, and is inconclusive.
The tip of the regenerate grows more rapidly than the rest; no elongation occurs at its cranial aspect.
Anguis , even when kept at 27°C, regenerates its tail very slowly, the best performance observed being a new tail of 5 mm after 14 weeks. The longest natural regenerate seen (16 mm) may have taken several years to produce in the wild.
The histological features of regeneration in Anguis are basically similar to those in other lizards. The new osteoderms are formed entirely in the subepidermal tissues but have a regular relationship with the scales. Some nerve fibres are regenerated with the ependymal tube.
The scales on the lizard's regenerating tail develop in a different manner from those in the lizard embryo and show suggestive resemblances to mammalian hairs.     

Inhibition of prolactin secretion by a direct effect of methysergide on the pituitary lactotrophs in the rat

Methysergide administered i.p. caused a dose dependent decrease of serum prolactin levels in rats of both sexes bearing large bilateral electrolytic lesions in the median eminence. This prolactin release inhibiting action of methysergide was prevented by pretreatment of the animals with dopamine receptor blockers pimozide or spiroperidol, which by themselves had no effect on serum prolactin levels. Similar results were observed when the dopamine receptor agonist piribedil was used instead of methysergide. It is concluded that methysergide is capable of inhibiting prolactin secretion by activation of dopamine receptors of the pituitary lactotrophs.     

Blood vessel formation during tail regeneration in the leopard gecko (Eublepharis macularius): The blastema is not avascular

Unique among amniotes, many lizards are able to self‐detach (autotomize) their tail and then regenerate a replacement. Tail regeneration involves the formation of a blastema, an accumulation of proliferating cells at the site of autotomy. Over time, cells of the blastema give rise to most of the tissues in the replacement tail. In non‐amniotes capable of regenerating (such as urodeles and some teleost fish), the blastema is reported to be essentially avascular until tissue differentiation takes place. For tail regenerating lizards less is known. Here, we investigate neovascularization during tail regeneration in the leopard gecko (Eublepharis macularius ). We demonstrate that the gecko tail blastema is not an avascular structure. Beginning with the onset of regenerative outgrowth, structurally mature (mural cell supported) blood vessels are found within the blastema. Although the pattern of blood vessel distribution in the regenerate tail differs from that of the original, a hierarchical network is established, with vessels of varying luminal diameters and wall thicknesses. Using immunostaining, we determine that blastema outgrowth and tissue differentiation is characterized by a dynamic interplay between the pro‐angiogenic protein vascular endothelial growth factor (VEGF) and the anti‐angiogenic protein thrombospondin‐1 (TSP‐1). VEGF‐expression is initially widespread, but diminishes as tissues differentiate. In contrast, TSP‐1 expression is initially restricted but becomes more abundant as VEGF‐expression wanes. We predict that variation in the neovascular response observed between different regeneration‐competent species likely relates to the volume of the blastema. J. Morphol. 278:380–389, 2017. © 2017 Wiley Periodicals, Inc.     

Prolactin-dependent seasonal changes in pelage: role of the pineal gland and dopamine.

The Siberian hamster displays seasonal changes in pelage that are dependent upon fluctuations in circulating prolactin levels. Pinealectomy prevented the decrease in serum prolactin and molt to the winter pelage displayed by castrated males housed under a short-day photoperiod. A dopaminergic antagonist, pimozide, enhanced prolactin levels in both pinealectomized and sham-operated animals under both long and short photoperiods. In the short-day animals, this effect of pimozide was associated with a prevention of the development of winter pelage. These results indicate that seasonal prolactin levels and related pelage changes are dependent upon the integrity of the pineal gland. However, basal prolactin levels under different photoperiod conditions appear to be only partly regulated by the actions of the dopaminergic system.     

Circadian participation in the photoregulation of testis activity and prolactin secretion in the mink, a short-day breeder

It has been demonstrated that an endogenous mechanism is involved in photoperiodic time measurement in the mink, a short-day-breeding mannal. A study of testicular activity (testicular volume, plasma testosterone concentration) and plasma prolactin level was carried out in sexually resting minks (the experiment began in November). Groups of minks were kept in the natural photoperiod or subjected to different resonance light-dark (LD) cycles (LD 4:8, LD 4:20, LD 4:32, LD 4:44); an additional group of animals was reared in an ahemeral photoperiod (LD 4:16). A rapid increase of testicular activity was observed in control animals or those kept in LD 4:20 (T 24) and LD 4:44 (T 48). In the other groups of animals, those kept in LD 4:8 (T 12), LD 4:32 (T 36), and LD 4:16 (T 20), testicular function remained at rest. Prolactin secretion was, in contrast, stimulated in the groups kept in LD 4:8 (T 12). LD 4:32 (T 36), and LD 4:16 (T 20), and remained low in the groups kept in LD 4:20 (T 24) and LD 4:44 (T 48). These results show that the effects of the different photoperiodic regimens do not depend on the duration of the photophase, but rather on the period of the LD cycles. The LD cycles that allow an increase of testicular function are those that are inhibitory to reproduction in birds and long-day-breeding mammals. To explain these results, it is suggested that in the mink exposure to light during the circadian photosensitive phase induces inhibition of testicular activity and stimulation of prolactin secretion. To explain the opposite effects of a single photoperiod on testicular function and secretion of prolactin, the hypothesis has been advanced that, in the mink, long days might simultaneously inhibit hypothalamic luteinizing-hormone-releasing hormone (LH-RH) activity and prolactin-inhibiting factor (PIF) activity.     

Synergetic effect of pimozide and thyrotropin releasing hormone on prolactin and thyrotropin release during the drying off of ewes

The effect of pimozide and/or TRH was investigated on plasma prolactin, thyrotropin, T₄ and T₃ and udder distension in 38 ewes during drying off by feed restriction. The effect of daily injections of 2 mg pimozide (s.c.), combined or not with TRH stimulation (200 μg, i.v.) on three different days of the drying off period was examined. Blood samples were taken twice daily in each group for 9 days, while blood sampling on the days of TRH injection was also performed at 0, 15, 30 min, and 1, 2 and 4 h post-injection. Plasma was assayed for PRL, TSH, T₄ and T₃ levels. Udder distension and mastitis incidence were recorded at the end of the drying off period. TRH and pimozide both resulted in elevated plasma PRL levels and acted in a synergetic way. Udder distension and the incidence of mastitis was only influenced by pimozide. The TSH as well as the T₃ response to TRH was increased in ewes under a continuous influence of pimozide and T₃ peaks following TRH injection occurred earlier than T₄ peaks. The higher effect of pimozide upon TRH stimulated PRL and TSH release at day 8 compared to days 0 and 3 indicates a progressive involvement of dopamine on the inhibition of PRL and the sensitivity of the thyrotrophs to TRH during drying off.     

Some functional aspects of canine corticotrophs

To examine the regulation and functional significance of canine pituitary pars intermedia corticotrophs, ACTH and cortisol responses to CRF were studied in healthy dogs before and after treatment with dexamethasone. In addition the effects of the dopamine agonist bromocriptine and the dopamine antagonist pimozide were investigated. In the latter two instances prolactin concentrations were also measured. Finally the pituitaries were studied immunocytochemically for ACTH and alpha-MSH. No response of ACTH or cortisol to bromocriptine was observed. Pimozide caused a slight rise in ACTH levels in some dogs. However, prolactin levels significantly decreased with bromocriptine and increased with pimozide. Injection of synthetic ovine CRF to dogs was followed by sharp increases in ACTH and cortisol values. These responses were obliterated by prior treatment with dexamethasone. In 1 of 4 dogs given dexamethasone before euthanasia, there were few pars distalis cells with ACTH(1-24) immunopositivity, although persistence of ACTH(1-24) reaction was noted within cells of the pars intermedia. The results indicate that none of the CRF-induced ACTH secretion in dogs is derived from pars intermedia corticotrophs. Dosages of bromocriptine and pimozide that clearly alter prolactin secretion do not consistently affect ACTH levels.     

Blastema cell proliferation in vitro: effects of limb amputation on the mitogenic activity of spinal cord extracts

Primary cultures of mesenchymal cells of axolotl limb blastemas provide a very sensitive in vitro bioassay for studying nerve dependence of newt regeneration. These cells can be stimulated by crude spinal cord extracts of non-amputated animals in a dose-dependent manner up to 60 micrograms protein/ml of culture medium; at this concentration the mitotic index is increased 4-fold. Spinal cord extracts of axolotls 14 days after forelimb amputation (i.e., late bud stage) are more efficient in stimulating blastema cell proliferation (+50%) than extracts of axolotls 7 days after forelimb amputation (i.e., early bud stage) or of axolotls without amputation. In a similar manner, spinal cord extracts of young axolotls 14 days after forelimb amputation, are more stimulatory than older axolotls 14 d after forelimb amputation which regenerate only a very small blastema during the same time. It appears that spinal cord mitogenic activity is enhanced after limb amputation, probably in correlation with blastema cell requirements for limb regeneration.     

Increased prolactin binding and morphological changes in the wound epithelium of regenerating limbs of Notophthalmus viridescens

Summary Distribution of prolactin has been examined in regenerating forelimbs from the newt Notophthalmus viridescens. Specific prolactin binding was demonstrated in homogenates of unamputated tissue, and of regenerating limbs at from 3 to 21 days postamputation. Labeled prolactin that was injected intraperitoneally into animals with one regenerating limb accumulated in the most distal portion of the regenerate at 7 and 14 days postamputation. Light microscopic autoradiography demonstrated that labeled prolactin was localized most heavily in the apical, outer layer of the wound epithelium. Scanning electron microscopy demonstrated that, in addition to changes in prolactin affinity following amputation, morphological changes occurred in the apical wound epithelium as well. Cell surfaces of the stump epidermis were characterized by periodic dispersion of papillae among a network of interconnecting structures 1–2 m across. By contrast, the surfaces of cells from the area in which labeled prolactin was found to localize most intensely were characterized by lack of papillae and, depending on the stage of regeneration, a pattern of microvilli and microplicae. These morphological alterations appear to reflect functional and biochemical differences between stump epidermis and wound epithelium.     

Differential effects of dopamine antagonists on prolactin secretion from cultured rat pituitary cells.

Various dopamine antagonists, including two novel non-neuroleptic drugs domperidone and halopemide, stimulated apomorphine-suppressed prolactin secretion from cultured rat pituitary cells. The potency of these drugs closely paralleled their rank-order in displacing $\text{in vitro}$ H³-haloperidol binding in rat striatum reported by others (10). Concentration-effect curves were parallel except those of pimozide and clopimozide which were biphasic : prolactin secretion was stimulated at low concentrations but depressed at concentrations above 25nM. When added alone, pimozide and clopimozide, but none of the other drugs tested, also depressed prolactin secretion. The present findings indicate that prolactin secretion from cultured pituitary cells may provide an $\text{in vitro}$ test system suitable to differentiate antagonists of dopamine receptors and possibly to distinguish pure from partial antagonists.     

Testis and epididymis of the Indian wall lizard (Hemidactylus flaviviridis): effects of flutamide on FSH and testosterone influenced spermatogenesis, Leydig cell, and epididymis

To determine the separate spermatogenic actions of FSH and testosterone, adult male lizards Hemidactylus flaviviridis with recrudescent testes were administered the non-steroidal antiandrogen flutamide either alone or in combination with FSH or testosterone, and the histology and histochemistry of the testes and ductus epididymides were studied. Flutamide-treated animals displayed a marked hypertrophy of Leydig cells. A few spermatids were also seen in testis of more than half the animals treated with flutamide. Flutamide also produced a significant increase of primary spermatocytes; no spermatids were observed in controls. A significant inhibition of spermatogenesis was noted in lizards treated either with testosterone alone or in combination with flutamide. Ovine FSH treatment caused a significant stimulation of spermatogenesis, as indicated by the increase of primary and secondary spermatocytes and the transformation of secondary spermatocytes into spermatids or, in a few cases, into spermatozoa. A considerable depletion of sudanophilic lipid and moderate delta 5-3 beta-hydroxysteroid dehydrogenase activity was noted in the Leydig cells of FSH-treated animals indicating enhanced steroidogenesis. Similar results were obtained when lizards were treated with flutamide + FSH. The effects of simultaneous treatment of flutamide with FSH or testosterone on ductus epididymidis revealed that flutamide markedly inhibited the epithelial cell height and lumen diameter with a loss of luminal content when compared to FSH or testosterone-treated lizards.     

Effects of pimozide and domperidone administration on prolactin levels in neonatally estrogenized female rats

The effects of two dopaminergic blockers, pimozide and domperidone, on the prolactin secretion were investigated in adult female rats treated neonatally with estrogens (100 micrograms of estradiol benzoate s.c. on day 1). These rats showed hyperprolactinemia (556 micrograms/l vs 57.7 in oil-injected) and treatment with pimozide or domperidone failed to increase prolactin levels in the adult age. These results suggest that the hyperprolactinemia in neonatally estrogenized female rats is produced by loss of the dopaminergic inhibition on prolactin secretion, so that the pharmacological blockade of dopaminergic receptors is uneffective. The dopamine levels in hypothalamus were similar in control and estrogenized females suggesting that failure in dopaminergic inhibition is due to a decrease in dopamine secretion to portal vessels.     

Increased ability of apomorphine to reduce serum concentrations of prolactin in rats treated chronically with alpha-methyltyrosine

Serum concentrations of prolactin were significantly increased in rats for up to 9 hours after a single i.p. injection of α-methyltyrosine (50 mg/kg); apomorphine caused a dose-dependent reduction of the elevated prolactin concentrations. Doses of apomorphine required to reduce serum prolactin concentrations.were lower in animals previously injected with α-methyltyrosine three times a day for 10 days than in animals which received a single injection of α-methyltyrosine. These results suggest that chronic disruption of the normal release of dopamine from tuberoinfundibular neurons leads to the development of increased sensitivity of dopamine receptors involved with the inhibition of prolactin release from the anterior pituitary.     

Interruption of the rat circadian clock by short light-dark cycles

Ninety male Sprague-Dawley rats were exposed to 1:1-h light-dark (LD1:1) cycles for 50-90 days, and then they were released into constant darkness (DD). During LD1:1 cycles, behavioral rhythms were gradually disintegrated, and circadian rhythms of locomotor activity, drinking, and urine 6-sulfatoxymelatonin excretion were eventually abolished. After release into DD, 44 (49%) rats showed arrhythmic behavior for >10 days. Seven (8%) animals that remained arrhythmic for >50 days in DD were exposed to brief light pulses or 12:12-h light-dark cycles, and then they restored their circadian rhythms. These results indicate that the circadian clock was stopped, at least functionally, by LD1:1 cycles and was restarted by subsequent light stimulation.     

Levels of cyclic GMP and cyclic AMP in regenerating forelimbs of adult newts following denervation

The effect of short-term denervation (0, 12, 24, and 72 hours) on the levels of cyclic 3'5'-guanosine monophosphate (cGMP) and cyclic 3'5'-adenosine monophosphate (cAMP) in adult newt (Notophthalmus viridescens) forelimbs at 15, 22, and 35 days of regeneration was investigated. Regenerate blastema and stump cyclic nucleotide levels were compared with those of the contralateral intact forelimb and hindlimb, with levels in the normally regenerating blastema, and with levels measured in the forelimbs of intact, nonoperated animals. Variations in cyclic nucleotide levels occurred according to regeneration stage and tissue type. Changes in level were noted immediately upon denervation and subsequently at other sample times in all regenerate and control series. Parallel fluctuations occurred in regenerate stump and contralateral intact forelimbs. Our results from nonamputated denervated and sham-denervated animals indicate that short-term, denervation-associated cyclic nucleotide fluctuations cannot be attributed solely to the loss of innervation.