Induction of photosensitivity in neonatal rat pineal gland

Abstract

Pineal glands removed from neonatal rats at 5, 7, and 9 days of age and explanted into short-term culture, synthesized melatonin when stimulated with norepinephrine (NE); their melatonin synthesis could not be suppressed with bright white light. Dispersed pineal cell cultures or pineal explants prepared from 1-day-old neonates and held in culture for 7 or 9 days also synthesized melatonin when stimulated with NE, but in these cases melatonin synthesis was significantly suppressed by light, demonstrating that the pineals had become photosensitive while in culture. The development of photosensitivity in culture could be partially or completely abolished by the continuous presence of 1 or 10 microm of NE in the culture medium. The pineals of all nonmammalian vertebrates are photoreceptive, whereas those of mammals do not normally respond to light. We hypothesize that a mechanism to suppress pineal photosensitivity by using NE released from sympathetic nerve endings evolved early in the history of mammals.

Figure 1

Melatonin levels ± SEM measured in dispersed pineal cell cultures (n = 10–12 cultures for each day). Cultures were obtained from postnatal day 1 rats and were maintained for 3, 5, 7, 9, 11, and 18 days before stimulation with NE. Melatonin levels were measured at the end of a 6-h period of stimulation with 10 μM of NE.

Figure 2

Photomicrograph showing the expression of AA-NAT mRNA in pineal cells after 7 days in dispersed cell culture (magnification ×63). (A) Antisense probe localized message in isolated cells that displayed neuron-like morphology with numerous fine neuritic processes; (B) Sense probe (control) showed no significant labeling.

Figure 3

Melatonin levels ± SEM measured in dispersed pineal cells cultured for 5, 7, and 9 days (n = 20–22 cultures for each treatment). Melatonin levels were measured at the end of a 6-h period of stimulation with 10 μM NE, during which the cells were exposed either to light (open bars) or dark (solid bars). ** denotes P < 0.01.

Figure 4

Melatonin levels ± SEM measured in pineal explants cultured for 5, 7, and 9 days (n = 19 cultures for each treatment). Melatonin levels were measured at the end of a 6-h period of stimulation with 10 μM NE, during which the explants were exposed either to light (open bars) or dark (solid bars). * denotes P < 0.05; ** P < 0.01.

Figure 5

Melatonin levels ± SEM measured in pineal explants obtained from neonatal rats at 5, 7, and 9 days of age (n = five cultures for each treatment). Melatonin levels were measured at the end of a 6-h period of stimulation with 10 μM NE, during which the explants were exposed either to light (open bars) or dark (solid bars). There were no significant differences between the light- and dark-exposed cultures.

Figure 6

Melatonin levels ± SEM measured in dispersed pineal cells cultured for 5, 7, and 9 days in medium containing (A) 1 μM NE and (B) 10 μM NE in fresh medium (see Materials and Methods; n = 16 cultures for each treatment). Melatonin levels in both cases were measured at the end of a 6-h period of stimulation with 10 μM NE, during which the cells were exposed either to light (open bars) or dark (solid bars). * denotes P < 0.05.