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A vinblastine sensitive high affinity choline uptake system
1Department of Physiology and Biophysics, University of Illinois, Chicago 60680.
Insights
The Limulus cardiac ganglion
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- The Limulus cardiac ganglion possesses a high-affinity choline uptake system (HAChUS).
- Cholinergic functions are described in other Limulus ganglia, but their HAChUS is unaffected by vinblastine.
- The role of HAChUS in the cardiac ganglion is not fully understood.
Purpose of the Study:
- To investigate the function of the Limulus cardiac ganglion HAChUS.
- To determine the specific role of choline uptake in cardiac ganglion physiology.
- To explore the relationship between choline uptake and membrane phospholipid synthesis.
Main Methods:
- Inhibition studies using vinblastine to assess HAChUS activity.
- Pulse-chase experiments to track radiolabeled choline incorporation.
- Enzymatic treatment (phospholipase C) to analyze radiolabeled compounds.
- Pharmacological manipulation with 5-hydroxytryptamine (5-HT) to study functional effects.
Main Results:
- Vinblastine significantly inhibited HAChUS in the cardiac ganglion in a dose-dependent manner.
- Radiolabel from [3H]choline increased in the pellet fraction post-incubation, and this increase was abolished by vinblastine.
- Phospholipase C treatment solubilized 72% of the radiolabel from the pellet fraction.
- 5-HT selectively reduced high-affinity choline uptake in the cardiac ganglion.
Conclusions:
- The Limulus cardiac ganglion HAChUS likely supplies choline for membrane phospholipid synthesis.
- This phospholipid synthesis may be linked to synaptic vesicle turnover.
- The cardiac ganglion's HAChUS has a distinct function compared to other Limulus ganglia.
Abstract:
1. The Limulus cardiac ganglion high affinity choline uptake system (HAChUS) was inhibited 40, 51 and 64% following pre-exposure to 10, 100 and 500 microM vinblastine, respectively. 2. In contrast, high affinity uptake of choline in the Limulus corpora pedunculata and abdominal ganglia, tissues in which a cholinergic function has been described, were unaffected. 3. In pulse-chase experiments, the cardiac ganglion was incubated in 0.1 microM [3H]choline for 60 min and then switched to an incubation medium containing 1 mM unlabelled choline for varying periods of time. 4. Under these conditions, a 3-fold increase of radiolabel above basal level was measured in the pellet fraction within 2 hr of post-labelling incubation. 5. Prior exposure of the ganglion to 500 microM vinblastine completely eliminated this increase of radioactivity in the pellet fraction. 6. Treatment of the radiolabelled pellet fraction with phospholipase C resulted in the solubilization of 72% of the radiolabel. 7. Ten (10) microM 5-hydroxytryptamine (5-HT), a concentration previously shown to inhibit spontaneous electrical activity within the cardiac ganglion, resulted in a 40% decrease in high affinity choline uptake in this tissue selectively. 8. These results are consistent with the view that a probable role of the Limulus cardiac ganglion HAChUS is the supply of choline subserving the synthesis of membrane phospholipid. 9. It is further speculated that this membrane phospholipid synthesis may be associated with synaptic vesicle turnover.