Development of enzyme activities and a circadian rhythm in pineal gland serotonin: evidence for a nonretinal pathway of light to the pineal gland of newborn rats.

The pineal gland of rats is markedly affected by exposure of the animals to varying periods of light and darkness. After maintenance of adult rats in continuous lighting, there is a significant decrease in pineal gland weight' and in the enzymatic capacity of the pineal gland to synthesize melatonin,2 and an increase in the activity of the serotonin-synthesizing enzyme, 5-hydroxytryptophan decarboxylase, in the pineal gland.3-5 These effects of light exposure are abolished by bilateral orbital enucleation.3-7 Circadian, or 24-hr rhythms, have been demonstrated in the content of serotonin in the rat pineal gland8 and in the activity of the melatonin-forming enzyme.9 The pineal content of serotonin is maximal at about 1 P.M.(lights on from 5 A.M. to 7 P.M.), and declines rapidly after the lights are turned off to reach a trough at about 11 p.M.8 We found'0 11 that this rhythm appears to be endogenous, since it persists in blinded animals and in rats kept in continuous darkness for up to 2 weeks. However, the nocturnal decline in pineal serotonin content can be prevented if illumination is extended an additional 4 hr to 11 p.M.8 11 Additional light exposure does not prevent the nocturnal decline in pineal serotonin in adult rats subjected to bilateral orbital enucleation, indicating that this effect required intact retinae." Intact retinae appear to be required for lighting information to influence most endocrine'2' 13 and circadian systems in mammals."'14 However, Ganong et al."5 have found that measurable amounts of light can penetrate the skull to the brains of mammals, without the intervention of the eyes. Other workers have obtained evidence suggesting that light can directly affect hypothalamic neurons-in the duck'6 and rat.'7 The present work was undertaken to study the development of the circadian rhythm in pineal gland serotonin content in newborn rats and its control by environmental lighting. Experiments were designed to examine the possibility that lighting information might influence the pineal gland by an extraretinal route in newborn rats. Methods.-Sprague-Dawley rats, both male and female, of varying ages were maintained with their mothers under diurnal lighting conditions in clear plastic cages at a constant temperature of 250C for at least 3 days prior to experimental treatment. An overhead fluorescent lamp provided about 110-150 ft-c of illumination at the level of the cages. Unless otherwise noted, lights were kept on from 5 A.M. to 7 P.M. daily. Rats were killed by anesthesia with chloroform at 1 P.M. and 11 P.M. Pineal glands were removed immediately, placed on paper towels impregnated with cold, isotonic saline, and weighed on a 25-mg Roller Smith balance. Two pineal glands were used for serotonin assay. After weighing, the pineal glands were homogenized in 0.5 ml of ice-cold 0.01 N hydrochloric acid with a conical ground-glass homogenizer and frozen. Serotonin assays were performed on the following day by the method of Snyder et al. 18