Cholinergic And Non-Cholinergic Afferents Of The Caudolateral Parabrachial Nucleus: A Role In The Long-Term Enhancement Of Rapid Eye Movement Sleep

Abstract

A single microinjection of the cholinergic agonist carbachol into the feline caudolateral parabrachial nucleus produces an immediate increase in state-independent ipsilateral ponto-geniculo-occipital waves, followed by a long-term rapid eye movement sleep enhancement lasting 7–10 days. Using retrogradely-transported fluorescent carbachol-conjugated nanospheres and choline acetyltransferase immunohistochemistry, afferent projections to this injection site for long-term rapid eye movement sleep enhancement were mapped and quantified. Six regions in the brain stem contained retrogradely-labelled cells: the raphe nuclei, locus coeruleus, laterodorsal tegmental nucleus, pedunculopontine tegmental nucleus, parabrachial nucleus, and the pontine reticular formation. The retrogradely-labelled (rhodamine+) cells in the pontine reticular formation and pedunculopontine tegmental nucleus contributed the predominant input to the parabrachial nucleus injection site (34.3 ± 5.3% and 28.4 ± 5.6%, respectively), compared to the laterodorsal tegmental nucleus (5.8 ± 3.8%), parabrachial nucleus (13.5 ± 3.1%), raphe nuclei (12.9 ± 2.7%), and locus coeruleus (5.1 ± 2.4%). By comparison with findings of afferent input to the induction site for short-latency rapid eye movement sleep in the anterodorsal pontine reticular formation, the parabrachial nucleus injection site is characterized by a similar proportion of afferents, except that the raphe nuclei were found to provide more than a two-fold greater input. Retrogradely-labelled neurons quantified in these nuclear regions consisted of 21.5% double-labelled (rhodamine+/choline acetyltransferase+) cholinergic and 78.5% noncholinergic (rhodamine+/choline acetyltransferase ) cells. The pedunculopontine tegmental nucleus contributed the predominant (51.7 ± 8.2%) cholinergic input, compared to laterodorsal tegmental nucleus (20.7 ± 10.2%), parabrachial nucleus (23.1 ± 7.5%), and pontine reticular formation (4.4± 2.1%). A comparative analysis of the total retrogradely-labelled cells within each nuclear region which were also double-labelled showed the highest proportion in the laterodorsal tegmental nucleus (76.2 ± 7.5%) compared to pedunculopontine tegmental nucleus (39.4 ± 3.6%), parabrachial nucleus (37.3 ± 2.8%), and pont±he reticular formation (3.2 ± 2.1%). These data indicate that while pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus neurons exert a powerful cholinergic influence on the injection site for long-term rapid eye movement enhancement, a major component of the afferent circuitry is non-cholinergic. Since the non-cholinergic input includes contributions from the locus coeruleus and raphe nuclei, it is probable that the caudolateral parabrachial nucleus contains cholinergic and aminergic afferent systems that participate in the long-term enhancement of rapid eye movement sleep.