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Date of Award

1984

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Medical Sciences

Supervisor

Dr. M.P. Rathbone

Abstract

The addition of dopamine to homogenates or slice preparations from rat caudate nucleus results in a 100-150% increase in tissue cAMP levels. Since protein kinases are the only known physiological receptors for cAMP, it has been hypothesized that the response of the postsynaptic neurons to dopamine may be mediated in part through the phosphorylation of specific membrane proteins. This study examines the effects of dopamine on cAMP and its associated protein kinase in caudate nucleus.

Caudate tissue contains a Type II protein kinase that is activated by cAMP, and that catalyzes the phosphorylation of several synaptic membrane proteins. Dopamine, however, did not appear to enhance the phosphorylation of these proteins either in broken cell or intact preparations, although increases in cAMP could be demonstrated under similar experimental conditions.

Destruction of postsynaptic neurons using the neurotoxin kainic acid significantly reduced cAMP-dependent protein kinase activity. Destruction of presynaptic nerve terminals, on the other hand, had no effect on the activity of this enzyme system. However, this procedure was associated with an increase in dopamine receptor sensitivity as measured by an increase in dopamine-mediated turning behaviour. This behavioural response was not abolished by intrastriatal administration of kainic acid, although this technique reduced dopamine-sensitive adenylate cyclase and cAMP-dependent protein phosphorylation by 80-90%.

The administration of chronic haloperidol also produced behavioural supersensitivity, as well as increases in the number of postsynaptic dopamine receptors, but this was not associated with comparable increases in the activity of dopamine-sensitive adenylate cyclase, or in cAMP-dependent protein phosphorylation. These studies therefore do not support a role for cAMP or cAMP-dependent protein kinases in the mediation of postsynaptic dopamine receptor supersensitivity.

Rat caudate nucleus also contains a number of proteins that are phosphorylated by Ca⁺⁺-dependent protein kinases. Substrates for calmodulin-dependent and independent protein kinases were identified and characterized on the basis of their solubilization properties and response to neuroleptic drugs. Dopamine enhanced the phosphorylation of two of these proteins in rat striatal slices. The effects of dopamine on protein phosphorylation could be distinguished from those of depolarizing agents such as KCl or veratridine, and were not mimicked by 8-bromo-cAMP.

The present studies demonstrate that increases in cAMP produced by dopamine have no measurable effect on cAMP-dependent protein phosphorylation. Although these data do not definitely preclude a role for cAMP-dependent protein kinases in the regulation of postsynaptic function, they indicate that the physiological relevance of dopamine-mediated increases in cAMP should be re-examined.

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