Dopaminergic transmission between identified neurons from the mollusk, Lymnaea stagnalis

Abstract

1. Dopaminergic transmission was investigated in the central nervous system (CNS) of the freshwater snail, Lymnaea stagnalis. 2. The giant pedal neuron, designated as right pedal dorsal one (RPeD1), makes chemical, monosynaptic connections with a number of identified follower cells in the CNS. Previous work has shown that RPeD1 is an interneuron and a important component of the Lymnaea respiratory central pattern generator. In this study, the hypothesis that RPeD1 uses dopamine as its neurotransmitter was tested by chromatographic, pharmacological, and electrophysiological methods. Characterization of RPeD1's transmitter pharmacology is essential to clearly understand its role in Lymnaea. 3. Earlier studies demonstrated that the soma of RPeD1 contains dopamine. This was quantitated in the present study by high-performance liquid chromatography (with electrochemical detection) of isolated RPeD1 somata and growth cones, which yielded 0.8 +/- 0.3 and 0.10 +/- 0.08 pmol of dopamine per soma and growth cone, respectively. 4. Bath or pressure application of dopamine to follower cells of RPeD1, in situ, mimicked the effects of RPeD1 stimulation. Dose-response curves were constructed for the excitatory effect of dopamine on follower cells, visceral dorsal two and three (VD2/3) (ED50 = 39 microM; Hill coefficient = 1.03), and the inhibitory effect of dopamine on follower cell, visceral dorsal four (ED50 = 33 microM; Hill coefficient = 0.92). 5. The following dopamine agonists (100 microM) were tested by bath application: 2-amino-6,7-dihydroxy-1,2,3,4-tetrahydronaphthalene (ADTN), apopmorphine, 2-bromo-alpha-ergocryptine, deoxyepinephrine (DE), mesulergine, (-) quinpirole, SKF 38393, and tyramine. Only the general dopamine agonists, ADTN and DE, mimicked RPeD1's effects on its follower cells. 6. When VD2/3 was isolated and plated in vitro, it maintained a depolarizing response to dopamine. This response was reduced by intracellular injection of the G-protein blocker, GDP-beta-S (2 mM in electrode). Similarly, incubation of VD2/3, in vitro for approximately 18 h, with pertussis toxin (PTX; 5 micrograms/ml), the G-protein inactivating exotoxin, also reduced the dopamine response. Injecting GDP or incubating in heat-inactivated PTX did not effect the response. 7. Several dopamine antagonists were used in an attempt to block RPeD1's synapses: chlorpromazine, ergonovine, fluphenazine, haloperidol, 6-hydroxydopamine, SCH 23390, (+/-) sulpiride, and tubocurarine. Only the D-2 dopamine receptor antagonist, (+/-) sulpiride, reversibly blocked synaptic transmission from RPeD1 to its follower cells. Both the (+) and the (-) enantiomer of sulpiride also antagonized synaptic transmission. A dose-inhibition curve for (+/-) sulpiride was constructed (IC50 = 47 microM).(ABSTRACT TRUNCATED AT 400 WORDS)