Cultured neurons from the thoracolumbar sympathetic chain of newborn mice are known to possess release-inhibiting ␣ 2 -autoreceptors. The present study was carried out in a search for release-modulating heteroreceptors on these neurons. Primary cultures were preincubated with [ 3 H]noradrenaline and then superfused and stimulated by single pulses, trains of 8 pulses at 100 Hz, or trains of 36 pulses at 3 Hz. The cholinergic agonist carbachol reduced the evoked overflow of tritium. Experiments with antagonists indicated that the inhibition was mediated by M 2 muscarinic receptors. The cannabinoid agonist WIN 55,212-2 reduced the evoked overflow of tritium through CB 1 receptors. Prostaglandin E 2 , sulprostone, and somatostatin also caused presynaptic inhibition. The inhibitory effects of carbachol, WIN 55,212-2, prostaglandin E 2 , and somatostatin were abolished (at the highest concentration of WIN 55,212-2 almost abolished) by pretreatment of the cultures with pertussis toxin (250 ng/ml). Several drugs, including the ␤ 2 -adrenoceptor agonist salbutamol, opioid receptor agonists, neuropeptide Y, angiotensin II, and bradykinin, failed to change the evoked overflow of tritium. These results demonstrate a distinct pattern of presynaptic inhibitory heteroreceptors, all coupled to pertussis toxin-sensitive G proteins. The lack of operation of several presynaptic receptors known to exist in adult mice in situ may be due to the age of the (newborn) donor animals or to the culture conditions. Key Words: Mouse sympathetic neuron cultures-Noradrenaline release -Presynaptic receptors-Muscarinic receptors-Cannabinoid receptors-Prostanoid receptors-Somatostatin receptors. J. Neurochem. 75, 2087Neurochem. 75, -2094Neurochem. 75, (2000.As in intact tissues, various receptors modulate the action potential-evoked release of noradrenaline in cultures of postganglionic sympathetic neurons from, for example, chicken and rats (for review, see Boehm and Huck, 1997). Neurons of mice, however, were not studied in this respect until we recently prepared primary cultures of thoracolumbar sympathetic ganglion cells from this species. The cells possessed release-inhibiting ␣ 2A/D -autoreceptors, but not release-modulating P2, A 1 , or A 2A receptors, which are alternative autoreceptors due to the cotransmitter role of ATP (Trendelenburg et al., 1999a, b). We have now carried out a search for releasemodulating heteroreceptors on these neurons. Most heteroreceptors ever observed were included (see Fuder and Muscholl, 1995;Powis and Bunn, 1995). The study concentrated on receptors modifying action potentialevoked release; receptors that, when activated, elicit exocytotic release of noradrenaline will be described in a subsequent report. Neuronal action potentials were triggered by electrical stimulation, and release of previously incorporated [ 3 H]noradrenaline was measured. MATERIALS AND METHODS Culturing and [ 3 H]noradrenaline releaseThese techniques were modified from our previous work (Trendelenburg et al., 1999a). Thora...

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The objective of the study was to clarify the postnatal development of the following transmitter release-modulating receptors of noradrenergic neurons in mice: alpha2-adrenoceptors, muscarinic, opioid and cannabinoid receptors (inhibitory), beta-adrenoceptors and receptors for angiotensin II and bradykinin (facilitatory). Wildtype (NMRI) and in some cases alpha2A/D-adrenoceptor-deficient mice aged 1 day (P1) or 8-16 weeks (adults) were used. Hippocampal and occipito-parietal cortex slices and sympathetically innervated tissues (atria and vas deferens) were preincubated with [3H]-noradrenaline and then superfused and stimulated electrically. Stimulation led to distinct increases in tritium efflux which were abolished by tetrodotoxin or removal of calcium. Concentration-response curves of appropriate agonists and in the case of alpha2-autoreceptors antagonists were determined. For beta-adrenoceptors and angiotensin receptors, the interaction of agonists with antagonists was also examined. Results demonstrate that alpha2A/D-autoreceptors operate already at P1 whereas nonalpha2A/D-autoreceptors, presumably alpha2C, develop later. Of the various heteroreceptors, those of brain noradrenergic neurons (OP3 and ORL1) modulate the release of [3H]-noradrenaline at least as effectively at P1 as in adults. Those of peripheral sympathetic neurons (muscarinic, probably mainly M2, OP1, OP2, OP3, CB1, AT1 and B1), in contrast, operate less effectively or not at all at P1, with one exception: beta2-adrenoceptors increase the release of [3H]-noradrenaline (atria) to the same extent, irrespective of age. Overall, results indicate that brain and peripheral noradrenergic neurons release their transmitter already shortly after birth. Presynaptic receptor mechanisms mature differentially in the brain and the periphery. Moreover, the various presynaptic receptors differ in their postnatal development and may play differential roles at different ages.