Conditioning the proboscis extension reflex of harnessed honeybees (Apis mellifera) is used to study the effect temporal spacing between successive conditioning trials has on memory. Retention is monitored at two long-term intervals corresponding to early (1 and 2 d after conditioning) and late long-term memory (3 and 4 d). The acquisition level is varied by using different conditioned stimuli (odors, mechanical stimulation, and temperature increase at the antenna), varying strengths of the unconditioned stimulus (sucrose), and various numbers of conditioning trials. How learning trials are spaced is the dominant factor both for acquisition and retention, and although longer intertrial intervals lead to better acquisition and higher retention, the level of acquisition per se does not determine the spacing effect on retention. Rather, spaced conditioning leads to higher memory consolidation both during acquisition and later, between the early and long-term memory phases. These consolidation processes can be selectively inhibited by blocking protein synthesis during acquisition.Learning trials distributed over time lead to better memory than learning trials squeezed into short periods of time. Jost (1897), who elaborated on the original findings by Ebbinghaus (1885), was the first to formulate a theory supposing a contraintuitive, even perplexing interrelation between short-term forgetting and long-term strengthening of memory. In his words, "Given equal associative strength, the older the memory trace at the time of learning repetition, the less forgetting over the long term." The "paradox of spaced practice" (Björk and Allen 1970) was assumed to lie in the fact that longer intervals between learning trials should lead to less memory on a trial-to-trial basis, but multiple units of less memory should finally lead to stronger long-term memory.The conceptual basis for interpreting the spacing effect is the notion of memory dynamics (James 1890; Müller and Pilzecker 1900;Squire 1987). Each learning trial is thought to initiate an intrinsic process of memory formation that leads to final memory by constructive (memory consolidation) and destructive (forgetting) processes. Studies of many species have shown that when training involves multiple trials, the time interval between trials is an important variable in the efficacy of accumulating training effects and the strength of retention (Carew et al. 1972;Fanselow and Tighe 1988;Tully et al. 1994;Spieler and Balota 1996;Kogan et al. 1997;Hermitte et al. 1999;Muzzio et al. 1999;Beck et al. 2000;Wu et al. 2001). The paradigms tested were taste aversion conditioning, fear conditioning, blink conditioning, olfactory aversion training, episodic priming, and learning nonsense syllables. In addition to humans, a wide range of animals was studied (Drosophila, the marine mollusks Aplysia and Hermissenda, the crab Chasmagnathus, rats, and rabbits). The dynamics and interdependence of the constructive and destructive (forgetting) memory processes, and, particularly, their relianc...

Extracellular recording were performed from mushroom body-extrinsic neurons while the animal was exposed to differential conditioning to two odors, the forward-paired conditioned stimulus (CSϩ; the odor that will be or has been paired with sucrose reward) and the unpaired CSϪ (the odor that will be or has been specifically unpaired with sucrose reward). A single neuron, the pedunculus-extrinsic neuron number 1 (PE1), was identified on the basis of its firing pattern, and other neurons were grouped together as non-PE1 neurons. PE1 reduces its response to CSϩ and does not change its response to CSϪafter learning. Most non-PE1 neurons do not change their responses during learning, but some decrease, and one neuron increases its response to CSϩ. PE1 receives inhibitory synaptic inputs, and neuroanatomical studies indicate closely attached GABA-immune reactive profiles originating at least partially from neurons of the protocerebral-calycal tract (PCT). Thus, either the associative reduction of odor responses originates within the PE1 via a long-term depression (LTD)-like mechanism, or PE1 receives stronger inhibition for the learned odor from the PCT neurons or from Kenyon cells. In any event, as the decreased firing of PE1 correlates with the increased probability of behavioral responses, our data suggest that the mushroom bodies exert general inhibition over sensory-motor connections, which relaxes selectively for learned stimuli.

Central interneurons exiting the alpha lobe of the mushroom bodies were studied with respect to their plasticity by electrically stimulating their presynaptic inputs, the Kenyon cells. Special attention was given to the analysis of a single, identified neuron, the PE1. Three stimulation protocols were tested: double pulses, tetanus (100·Hz for 1·s), and tetanus paired with intracellular deor hyper-polarization of the recorded cell. Double-pulse stimulations revealed short-term facilitation and depression, tuning the responses of these interneurons to frequencies in the range of 20-40·Hz. The tetanus may lead to augmentation of responses to test stimuli lasting for several minutes, or to depression followed by augmentation. Associative long-term potentiation (LTP) was induced in the PE1 neuron by pairing a presynaptic tetanus with depolarization. This is the first time that associative LTP has been found in an interneuron of the insect nervous system. These data are discussed in the context of spike tuning in the output of the mushroom body, and the potential role of associative LTP in olfactory learning. It is concluded that the honeybee mushroom body output neurons are likely to contribute to the formation of olfactory memory.

Degen et al. used a special radar system to track bees in flight. They displaced bees after a single orientation flight into either the explored or the unexplored area. Homing flights were faster and straighter if bees were released within the explored area. The authors conclude that bees used the ground structure for homeward guidance.

In the honeybee brain, two prominent tracts – the medial and the lateral antennal lobe tract – project from the primary olfactory center, the antennal lobes (ALs), to the central brain, the mushroom bodies (MBs), and the protocerebral lobe (PL). Intracellularly stained uniglomerular projection neurons were reconstructed, registered to the 3D honeybee standard brain atlas, and then used to derive the spatial properties and quantitative morphology of the neurons of both tracts. We evaluated putative synaptic contacts of projection neurons (PNs) using confocal microscopy. Analysis of the patterns of axon terminals revealed a domain-like innervation within the MB lip neuropil. PNs of the lateral tract arborized more sparsely within the lips and exhibited fewer synaptic boutons, while medial tract neurons occupied broader regions in the MB calyces and the PL. Our data show that uPNs from the medial and lateral tract innervate both the core and the cortex of the ipsilateral MB lip but differ in their innervation patterns in these regions. In the mushroombody neuropil collar we found evidence for ALT boutons suggesting the collar as a multi modal input site including olfactory input similar to lip and basal ring. In addition, our data support the conclusion drawn in previous studies that reciprocal synapses exist between PNs, octopaminergic-, and GABAergic cells in the MB calyces. For the first time, we found evidence for connections between both tracts within the AL.