Effects of adrenergic agonists and antagonists on the numbers of synaptic ribbons in the rat pineal gland.

In the pineal gland numbers of synaptic ribbons (SR) undergo day/night changes which parallel the rhythm of melatonin synthesis. Since pineal biosynthetic activity is controlled by activation of adrenoreceptors, we investigated the effects of adrenergic agonists and antagonists on pineal synaptic ribbon numbers and N-acetyltransferase (NAT) activity, the key enzyme of melatonin synthesis in rats. In vivo application of the beta-adrenergic antagonist propranolol decreased melatonin synthesis when given during the dark phase but did not affect SR numbers. Treatment during daytime with the beta-adrenergic agonist isoproterenol increased pineal NAT activity whereas SR numbers did not change. Norepinephrine stimulated NAT activity in vitro in a dose-dependent manner, but did not elevate SR numbers. Incubation with an analog of the second messenger cyclic adenosine monophosphate increased both NAT activity and SR numbers. These results suggest that the beta-adrenergic system does not play a decisive role in the regulation of the nocturnal increase in SR numbers observed in the rat pineal gland.

Comparison of radioimmunoprecipitation assays for the detection of human anti-tumor virus antibodies.

The demonstration of human antibodies reactive in radioimmunoprecipitation assays (RIAs) with primate tumor virus (oncornavirus) antigen has implications for a possible previously published negative findings and led to considerable scientific controversy. We feel much of the discrepancy may be of methodological origin. An attempt is therefore made in this communication to resolve these apparent discrepancies by comparing various published parameters of the RIAs used in the search for human antibodies reactive with oncornavirus antigens.

One millisecond of light suffices to suppress nighttime pineal melatonin synthesis in rats.

The effect of a single high-intensity light pulse with a duration of 1 ms on nighttime pineal activity of male Sprague-Dawley rats was investigated. 10 minutes after light exposure pineal N-actyltransferase activity and melatonin content were significantly reduced. These results show that the rat pineal is capable of responding to very short light flashes of high intensity.

Response of pineal serotonin N-acetyltransferase activity in male guinea pigs exposed to light pulses at night.

Serotonin N-acetyltransferase (NAT), which is crucial for the formation of melatonin, undergoes a typical day/night rhythm in the pineal gland with low levels during daytime and high levels at night. Short pulses of light given at night have been shown to rapidly depress NAT activity in some species, but not in others, the reasons for this difference being unclear. As diurnality and nocturnality of the experimental animals may play a role and since diurnally active animals have been little investigated in this respect, in the present study the diurnally active guinea pig was investigated. Male guinea pigs kept under a lighting regimen of LD 12:12 (lights off at 1700 hrs) were killed between 1200 or 1300 hrs and between 0000 and 0200 hrs, at night in the dark or after exposure to 10 or 45 min of light. The results obtained show that the day/night difference of NAT activity is about 2-fold. 10 min or 45 min of light given at night significantly depress pineal NAT activity. Re-exposure to darkness for 1 hr of animals previously given light for 10 min leads to restoration of NAT activity. These findings together with data from the literature suggest that it does not appear to be the activity pattern (diurnality versus nocturnality) of an animal nor the amplitude of the day/night difference of pineal NAT activity that account for the suppressibility or non-suppressibility of pineal NAT activity by light at night.

Cell aggregation and sexual differentiation in pairs of aggregation-deficient mutants of Dictyostelium discoideum.

A diffusible aggregation-stimulating factor (ASF) is released from a series of aggregation-deficient mutants. Biochemical markers indicate that these ASF-donor mutants are blocked at a later step of cell differentiation than an ASF-requiring mutant. ASF is able to bridge the initial block of differentiation in the latter mutant, such that development proceeds up to aggregation and even further. ASF is most probably neither identical with cyclic-AMP phosphodiesterase nor with an inhibitor of this enzyme which both are released from donor strains. In certain combinations of aggregation-deficient mutants, macrocysts, the sexual stages of Dictyostelium, are formed. Also, motile giant cells believed to be zygotes are observed in these mutant combinations. The gamone known to be released from one mating type and to induce macrocysts in the other, is probably not identical with ASF since this factor is produced by mutants derived from either one of both mating types.

Day/night serotonin levels in the pineal gland of male BALB/c mice with melatonin deficiency.

Studies from another laboratory have shown that several strains of laboratory mice have a genetic defect for melatonin synthesis. In non-deficient species, melatonin synthesis undergoes a typical, beta-adrenergically regulated day/night rhythm with low melatonin levels during daytime and high levels at night, the precursor serotonin showing an inverse behaviour. This study examines whether a day/night rhythm of pineal serotonin levels exists in melatonin-deficient male BALB/c mice. Mice kept under a lighting schedule of 12 h light (lights on at 07.00 h) and 12 h dark were killed at 13.00 and 01.00 h, respectively. Serotonin amounted to 12-15 ng/pineal and did not show regular day/night differences. Administration of the beta-adrenergic agonist, isoproterenol, which is known to affect melatonin synthesis in a number of species, was without effect on pineal serotonin levels. Melatonin and two of the melatonin-forming enzymes, serotonin N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) were below the detection limit in the pineal. It is concluded that in melatonin-deficient BALB/c mice, pineal serotonin synthesis is apparently intact. In BALB/c mice, serotonin synthesis and release do not appear to be directly or indirectly regulated by beta-adrenergic mechanisms.

The effects of short pulses of light at night on numbers of pineal "synaptic" ribbons and serotonin N-acetyltransferase activity in male Sprague-Dawley rats.

To characterize further the functionally enigmatic "synaptic" ribbons (SR) of the mammalian pineal gland and to study possible relationships to melatonin synthesis, in the present investigation rats were exposed to short pulses of light at night when both SR numbers and serotonin N-acetyltransferase (NAT) activity are high in comparison to day-time values. Male Sprague-Dawley rats were killed at 13:00 and 01:00 h, respectively, and at 01:10 and 02:00 h after exposure to light for 10 and 60 min, respectively. The pineals were rapidly taken out and cut sagittally in half. One half was processed for electron-microscopic quantitation of SR numbers and the other half for NAT determinations. It was found that both SR numbers and NAT activity decreased significantly when the animals were exposed to light at night. Although both parameters showed corresponding changes, there was no clear-cut correlation between SR numbers and NAT activity in individual animals within a group, except after exposure to light for 60 min when a positive correlation (R = 0.939; p less than 0.05) existed. After exposure to light the electron-lucent vesicles of the SR decreased in number, but the length of the SR was unchanged. These results show that numbers of pineal SR can be easily and quickly manipulated and that the presently used model may be ideal in studying the poorly understood mode in which degradation of SR occurs.

The immune response against the ASV-coded src-gene product in syngeneic mice.

The antigenicity of the avian sarcoma virus (ASV)-coded src-gene product pp60src, which is responsible for fibroblast transformation after ASV infection, has been investigated in STU mouse fibrosarcoma cell lines and the corresponding immune response in syngeneic mice has been determined. The development of effective anti-pp60src antibody titres depends on the mode and stie of injection of tumour cells and parallels tumour growth. It was found that mouse immunoglobulin heavy chains are unable to serve as substrate for the protein kinase activity of pp60src. Therefore, an indirect protein kinase absorption (PKA) test was initiated to demonstrate recognition of the protein kinase activity associated with the src-gene product. The availability of syngeneic mice and the corresponding ASV-transformed tumour cells should facilitate studies designed to elucidate the possible relationship between the cytoplasmic pp60src and ASV-induced tumour-specific surface antigens (TSSA), for example, by allowing the production of stable mouse hybridomas synthesizing antibodies specific for pp60src and TSSA.

Serotonin and melatonin contents in the pineal glands from different stocks and strains of laboratory rats.

In the present study the pineal gland was examined in 2 outbred stocks and 6 inbred strains of rats some of which were pigmented to varying degrees, to see whether inbreeding affects the variability and whether differences exist between albino and pigmented rats. The animals were kept under 12 h light: 12 h darkness (12 L:12 D) and killed 7 h after the onset of light and darkness, respectively. The parameters examined were pineal protein content, serotonin and melatonin levels and hydroxyindole-O-methyltransferase (HIOMT) activity. All the parameters examined revealed interstrain differences, independently of whether the data were expressed per pineal or per mg protein. The variation coefficients for the various parameters were relatively high. They were mostly smaller when the data were expressed per pineal rather than per mg protein. No striking differences existed between the variation coefficients in inbred and outbred rats. When pineal size and the melatonin-related parameters expressed per pineal were used to assess the melatonin-synthesizing capacity of the pineal glands, it was found that the outbred Wistar and Sprague-Dawley rats and the inbred LEWIS-derived (LEW/Han) rats, all of which were albinos, had the most active pineals. Intermediate activity was noted in the hooded E3/Han and BDE/Han and the albino BDII/Han rats. The smallest and least active pineals were found in the totally pigmented BN/Han and DA/Han rats. The results taken together show that different stocks and strains exhibit significant differences in pineal size and melatonin-forming capacity. Albino rats appear to have larger and more active pineals than pigmented rats.

The pineal complex in Roman high avoidance and Roman low avoidance rats.

Previous studies have shown that the pineal gland of Roman high avoidance (RHA/Verh) rats is larger than that of Roman low avoidance rats (RLA/Verh). In the present study measurement of enzyme activities (serotonin-N-acetyl-transferase, hydroxyindole-O-methyltransferase) revealed that pineals of RHA/Verh rats are twice as active in melatonin production than pineals of RLA/Verh rats. Indoleamine content was also higher in RHA/Verh rats, whereas noradrenaline content was the same in both lines. When values were expressed per mg protein, these differences disappeared except for N-acetyl-serotonin and noradrenaline which were higher or lower in RHA/Verh rats, respectively. Both lines had higher serum levels of melatonin during the dark phase than during the light phase. However, RHA/Verh rats had increased serum levels as compared to RLA/Verh rats during both day and night. Morphometric analysis of the deep and superficial part of the pineal complex revealed, that the volumes of both parts are enlarged in RHA/Verh rats. Electron microscopic studies of pineals collected during day- and nighttime showed higher numbers of synaptic ribbons per unit area in pineals of RHA/Verh rats. In pineals collected during June synaptic ribbons displayed a day/night rhythm in RHA/Verh rats only, whereas in glands of both lines collected during November no daily changes were found. These results show that closely related but divergently selected rat lines may differ in pineal ultrastructure and pineal function.