No abstract

The relationships among biochemical parameters of cholinergic metabolism in autonomic nerve terminals in the chick iris were determined throughout the life span of the animal, following in vitro incubation with [³H]choline (Ch). Choline accumulation, acetylcholine (ACh) content and ACh turnover increased steadily up to the adult stage (3 months – 1 year) and then declined in aged animals (5 years). These changes appeared to reflect the activity and the state of the cholinergic innervation of the iris. Other changes which occurred with age included a steady decline in the ratio of Ch to ACh, and a decrease in the concentration of endogenous Ch relative to [³H]Ch taken up from the medium. However, the ratio of [³H]Ch to [³H]ACh did not vary significantly with age from 10 days of incubation (d.i.) up to 5 years. Uptake of [³H]Ch and synthesis of [³H] ACh were both inhibited by low temperature (27°C), ouabain (1CH M), hemicholinium (HC-3) (5 x 10^–5M) and low Na⁺ at both 3 months and 5 years. Low temperature and ouabain, however, did not inhibit [³H]ACh formation to the same extent as HC-3 and low Na⁺. Cholinesterase inhibitors appeared to inhibit [³H]Ch uptake by increasing intracellular ACh, confirming previous reports. These observations can be taken to support our view that the characteristics of Ch uptake do not develop and age simultaneously in cholinergic endings. In conclusion, our results indicate that the turnover rate of ACh is slowed in aging autonomic nerve terminals, although the rate of synthesis of [³H]ACh from taken up [³H]Ch is unvaried. The present findings on Ch metabolism taken together with our previous results on ACh and Ch content, Ch uptake, and enzymatic activities confirm our view of the peripheral terminals of the autonomic neuron as a site of specific developmental changes and of selective vulnerability to aging processes.

Age-related changes in the activities of choline acetyltransferase and acetylcholinesterase in avian sympathetic ganglia, ciliary ganglia and iris have been determined throughout the posthatching life span. Kinetic characteristics of these enzymes have been analyzed in the nerve terminals in the iris from early embryonic stages to the latest periods of life. Enzyme activities and kinetics have been related to each other and also to the endogenous levels of acetylcholine. The regulation of the biosynthetic mechanism for acetylcholine through choline uptake and choline acetyltransferase activity in the developing nerve terminal have been examined. The activities of both enzymes in all tissues are reduced at later ages, which is referred to declines in V_max, as the K_m values do not vary significantly at any age. Declines with age are most pronounced in the iris, both in choline acetyltransferase activity and V_max and in acetylcholine levels, supporting our view of the peripheral autonomic nerve terminal as a major locus for the effects of aging on the peripheral nervous system.

Choline (Ch) uptake in the iris of the chick has been characterized from 4½ days of incubation to 1 year of age, in conjunction with an electron microscopic ultrastructural study of the innervation of the iris. Uptake has been shown to constitute an early and reliable biochemical marker for the presence of innervation in target organs. The characteristics of Ch uptake, including Na⁺ and temperature dependence and inhibitor sensitivity, change during development, and the V_max increases steadily while the K_m does not vary significantly during the period of study. The relationships among uptake and other cholinergic functions also change with development of the iris. Acetylcholine (ACh) and Ch uptake but not Ch levels appear to develop together, while cholinergic enzyme activities follow a developmental course more closely related to neurotransmission. The significance of these findings in relation to proposed regulatory relationships in cholinergic biosynthetic mechanisms is discussed.