BEHAVIOURAL BRAIN RESEARCH ELSEVIER BehaviouralBrain Research 81(1996) 215 222 , , , Research report Mecamylamine-induced impairment of acquisition and retrieval of olfactory conditioning in the honeybee Val~rie Cano Lozano, Elisabeth Bonnard, Monique Gauthier *, Daniel Richard Laboratoire de Neurobiologie et Comportement, Universit~ de Toulouse III, 118 route de Narbonne, 31062 Toulouse Cedex, France Received 6 June 1994; revised 18 March 1996; accepted 21 March 1996 Abstract Mecamylamine, a nicotinic receptor antagonist, was injected into the honeybee brain haemolymph. The effects of the drug were investigated on Pavlovian conditioning of the proboscis extension reflex. The conditioned response was acquired after a one-trial learning session, consisting of an olfactory-conditioned stimulus combined with a gustatory antennal unconditioned stimulus. The drug was injected at different times before or after the learning session in order to dissociate its effects on acquisition, consolidation and retrieval processes. The performance was evaluated in short-delayed recall tasks. To control potential effects on sensory-motor activity, the effects of the drug were also investigated on sensory processes (through olfactory and gustatory functions) and on motor processes of proboscis extension. The results of conditioning experiments showed that pretrial injection induced a decrease of retention performance 1 h after the learning trial. Mecamylamine injected 20 min after the learning session induced a time- dependent impairment of retention performance, as has been shown by the performance level registered from 10 to 80 min after injection. A 5-min post-trial injection had no effect on retention performance. Control experiments did not reveal any effect of mecamylamine on the response reflex of proboscis extension and on responsiveness to olfactory stimuli (geraniol, lavender and vaniUin). The absence of effects on sensory perception combined with the amnestic effect induced by pre- or late post-trial injections lead us to conclude that mecamylamine specifically impaired acquisition and retrieval processes. The involvement of nicotinic-like receptors in these processes is discussed. Keywords: Honeybee; Olfactory conditioning; Memory; Nicotinic receptor; Mecamylamine 1. Introduction In the honeybee, the olfactory conditioning of the proboscis extension reflex is a very useful paradigm to study the neurobiological basis of associative learning [22]. This Pavlovian procedure is based on the elicita- tion of the reflex by an initially non-effective olfactory stimulus. Data from the literature show that a single learning trial is sufficient to condition a high percentage of honeybees [4]. This procedure, based on a one-trial learning session, allows acquisition, storage and retrieval processes to be separated and manipulated [25]. The effects of biogenic amines on learning and memory processes have been tested with this procedure and intracranial injections of agonists and antagonists * Corresponding author. Fax: + 33 6155-8444; E-mail: gauthiem@cict.fr 0166-4328/96/$15.00 © 1996 Elsevier Science B.V. All rights reserved PII S0166-4328 (96)00055-1 revealed that monoamines are important modulators of memory formation and retrieval [28,29]. In parallel to monoamines, acetylcholine (ACh) is a widespread neurotransmitter in the central nervous system of insects [6,10]. In contrast to vertebrates, the major part of central cholinergic receptors exhibits nico- tinic properties, but muscarinic binding sites have also been identified [7,39]. In the central nervous system of the honeybee, acetylcholine is probably the less well- known neurotransmitter. Pharmacological investiga- tions have shown that nicotinic and muscarinic binding sites are present in the brain of the honeybee [1,20]. Moreover, Kreissl and Bicker [21] have shown that 0~-bungarotoxin (0t-BGT) binding sites in brain neurop- iles coincide with acetylcholinesterase (ACHE) staining, suggesting the presence of nicotinic receptors. It was recently shown in Kenyon cell culture that these recep- tors are functional [3]. The role of ACh in nervous function is yet not well