Norepinephrine release in the rat pineal gland: the input from the biological clock measured by in vivo microdialysis.

The sympathetic innervation of the rat pineal gland was investigated, measuring the norepinephrine (NE) release by on-line in vivo microdialysis. NE was assayed using an HPLC method with precolumn derivatization and fluorescence detection. Its high sensitivity and reliability made it very suitable to monitor the low levels of NE in the dialysates (12.5 fmol during nighttime, 3 fmol during daytime). To increase NE levels, the monoamine reuptake inhibitor cocaine was added to Ringer's solution at concentrations of 10(-6) and 10(-5) M. This resulted in increases of neurotransmitter output of 167 and 219%, respectively, but did not change the qualitative and/or quantitative outcome of other experiments. Perfusion with 10(-6) M tetrodotoxin for 1 h resulted in a decrease of the NE release by >80%, whereas perfusion with the alpha 2-receptor antagonist yohimbine caused a twofold increase. These results indicate that the NE release in the rat pineal was of neuronal origin and regulated by a negative feedback mechanism involving inhibitory presynaptic alpha 2-receptors. Long-term (i.e., 16 h) measurements are described, showing the circadian properties of NE release. A pronounced rhythm is reported, showing extremely sharp transitions between low daytime and high nighttime values. Increases and decreases are reported to occur within the duration of collecting one sample (20 min). For comparison, the rhythm of melatonin release was also recorded. The on and off switches of the sympathetic input correlated well with the circadian rhythm of melatonin release and can thus be considered as the primary clock signal, inducing the nightly production of melatonin.

Microdialysis reveals dynamics of coupling between noradrenaline release and melatonin secretion in conscious rats.

The coupling between noradrenergic innervation of the pineal gland and melatonin production was investigated. Previously, the development of a microdialysis technique was described which made it possible to study the noradrenaline (NA) input as well as the melatonin output with high time resolution. In the present study, we studied the effects of short-term changes in NA-release on melatonin secretion. A 1 min light pulse was applied around midnight and resulted in an immediate decrease of both NA and melatonin. While NA returned to basal levels in 40 min, melatonin did not reach the baseline within 2.5 h. This discrepancy in correlation between NA and melatonin indicates a rapid inactivation of N-acetyl-transferase (NAT), followed by a slow reactivation, possibly by de novo synthesis of NAT. During daytime, a perfusion with 60 mM potassium for 30 min, resulted in a rapid and short stimulation of NA release, which was not followed by an increase in melatonin production. This indicates that 30 min stimulation of NAT activation is not enough to increase the amount of melatonin produced. The combination of measuring NA input and melatonin output appears to be a valuable tool in studying the characteristics of pineal activity in great detail.