Melatonin receptor deficiency decreases and temporally shifts ecto-5'-nucleotidase mRNA levels in mouse prosencephalon.

Ecto-5'-nucleotidase (eN) is the major extracellular adenosine-producing ecto-enzyme in mouse brain. Via the production of adenosine, eN participates in many physiological and pathological processes, such as wakefulness, inflammation, nociception and neuroprotection. The mechanisms regulating the expression of eN are therefore of considerable neurobiological and clinical interest. Having previously described a modulatory effect of melatonin in the regulation of eN mRNA levels, we decided to analyze the melatonin receptor subtype involved in the regulation of eN mRNA levels by comparing eN mRNA patterns in melatonin-proficient transgenic mice lacking either the melatonin receptor subtype 1 (MT1 KO) or both melatonin receptor subtypes (MT1 and MT2; MT1/2 KO) with the corresponding melatonin-proficient wild-type (WT) controls. By means of radioactive in situ hybridization, eN mRNA levels were found to be diminished in both MT1 and MT1/2 KO mice compared with WT controls suggesting stimulatory impacts of melatonin receptors on eN mRNA levels. Whereas eN mRNA levels increased during the day and peaked at night in WT and MT1 KO mice, eN mRNA levels at night were reduced and the peak was shifted toward day-time in double MT1/2 KO mice. These data suggest that the MT2 receptor subtype may play a role in the temporal regulation of eN mRNA availability. Notably, day-time locomotor activity was significantly higher in MT1/2 KO compared with WT mice. Our results suggest melatoninergic signaling as an interface between the purinergic system and the circadian system.

Expression of ectonucleotidases in the prosencephalon of melatonin-proficient C3H and melatonin-deficient C57Bl mice: spatial distribution and time-dependent changes.

Extracellular purines (ATP, ADP, AMP and adenosine) are important signaling molecules in the CNS. Levels of extracellular purines are regulated by enzymes located at the cell surface referred to as ectonucleotidases. Time-dependent changes in their expression could profoundly influence the availability of extracellular purines and thereby purinergic signaling. Using radioactive in situ hybridization, we analyzed the mRNA distribution of the enzymes NTPDase1, -2 and -3 and ecto-5'-nucleotidase in the prosencephalon of two mouse strains: melatonin-proficient C3H and melatonin-deficient C57Bl. The mRNAs of these enzymes were localized to specific brain regions, such as hippocampus, striatum, medial habenula and ventromedial hypothalamus. NTPDase3 expression was more widely distributed than previously thought. All ectonucleotidases investigated revealed a prominent time-dependent expression pattern. In C3H, the mRNA expression of all four enzymes gradually increased during the day and peaked during the night. In contrast, in C57Bl, ecto-5'-nucleotidase expression peaked at the beginning of the day and gradually decreased to trough levels at night. Recording of locomotor activity revealed higher daytime activity of C57Bl than of C3H. Our results indicate that the expression of ectonucleotidases varies according to time and genotype and suggest that melatonin exerts modulatory effects associated with different regulations of purinergic signaling in the brain. These findings provide an important basis for further examination of the complexity of the purinergic system in the brain.