An increasing body of evidence indicates that macromolecules derived from target organs are essential for the survival of embryonic sympathetic neurons (SN) in culture. Whether these factors also influence the development of noradrenergic characteristics (i.e., uptake, storage, synthesis and release of norepinephrine (NE) is virtually unknown. An attractive possibility exists that survival and development of SN and their functional maturation may be controlled by discrete influences. The Principal Investigator (PI) has successfully attempted to measure one of the important functions of SN maintained in culture. It is possible to evoke Ca-dependent release of 3H-NE by electrical stimulation of SN maintained in culture. The potential advantages of the method are: 1) Release can be studied from essentially a pure population of SN; 2) SN can be obtained from embryos of different ages, allowing to study noradrenergic function in relation to development; 3) SN can be maintained in culture for long periods in the absence of non-neuronal cells and serum; 4) SN can be co-cultured with muscle cells, other types of neuronal and non-neuronal cells to study interactions between cells and their effect on noradrenergic properties. In preliminary experiments PI has studied release properties of embyronic SN, and they appear to be dramatically different from those of mature mammalian SN. A more intriguing observation is that cultured SN differed in their release properties even from their equivalents growing in the embryonic heart. For example, release evoked from cultured SN was not blocked by tetrodotoxin (TTX) or facilitated by tetraethylammonium (TEA). The release was not even under the control of presynaptic adrenergic modulation. In sharp contrast, release evoked from SN innvervating the embryonic heart was blocked and potentiated by TTX and TEA, respectively, and modulated by exogenous NE. Ca was essential for the release in both situations. The immediate objective is to establish noradrenergic characteristics of embryonic SN growing in culture and in the heart. The long-term objective is to establish whether the target organ plays a role in the development of noradrenergic characteristics of their neurons during synaptogenesis by examining noradrenergic properties in SN cultured under different conditions, particularly in the presence of heart cells. It is hoped that the proposed experiments will reveal that survival and development of SN and maturation of their functional properties are controlled by different signals.
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