Wj. Drijfhout et al., MICRODIALYSIS OF MELATONIN IN THE RAT PINEAL-GLAND - METHODOLOGY AND PHARMACOLOGICAL APPLICATIONS, Journal of neurochemistry, 61(3), 1993, pp. 936-942
The present study describes the development of a new technique to meas
ure melatonin contents in the pineal gland of freely moving rats, by m
eans of on-line microdialysis. The transcerebral cannula was modified,
and a sensitive assay of melatonin, using HPLC with fluorimetric dete
ction, was set up. With this system it is possible to monitor the mela
tonin levels on-line in the pineal gland during day- and nighttime. Th
e nightly increase in melatonin release was recorded. Tetrodotoxin had
an inhibitory effect on nighttime levels, whereas even high concentra
tions did not alter the daytime level. From this we conclude that neur
onal activity is necessary to synthesize melatonin and that during day
time no net neuronal activity is present. Melatonin levels could be gr
eatly enhanced by systemic administration of the beta-agonist isoprena
line (ISO). Also, local infusion of ISO or 8-bromoadenosine 3',5'-cycl
ic monophosphate, an analogue of the second messenger cyclic AMP, resu
lted in increased melatonin levels, demonstrating the presence of beta
-adrenergic receptors, coupled to a cyclic AMP-based second messenger
system, on the pineal gland. Injection of phenylephrine had no effect
on daytime levels. Only when administered during ISO-induced stimulati
on of melatonin release did it enhance this stimulated release. This p
roved the regulatory role of alpha1-receptors on pinealocytes. The met
hod presented is of special interest for investigating the innervation
of the pineal gland and the biochemical processes that regulate the b
iosynthesis of melatonin. Also, for studies on the diurnal rhythms of
melatonin release and factors that influence these rhythms in freely m
oving animals, this model will be of great value.