Steven Rose

How chicks make memories: the cellular cascade from c-fos to dendritic remodelling

Training chicks on a one-trial passive avoidance task results in a cellular cascade over the subsequent hours. Phosphorylation of the presynaptic phosphokinase C substrate B-50 is followed by immediate-early gene expression and increased synthesis of pre- and postsynaptic glycoproteins, increases in dendritic spine densities, synapse and synaptic vesicle numbers, and a prolonged increase in neuronal bursting. Many of these effects have been localized to two forebrain regions: the left intermediate medial hyperstriatum ventrale and the lobus parolfactorius. Pretraining lesions in the left intermediate medial hyperstriatum ventrale, or post-training lesions in the lobus parolfactorius result in amnesia. These and related results lead to models of memory storage based on multiple representation by way of synaptic stabilization through glycoprotein synaptic recognition molecules.

Corticosterone enhances long-term retention in one-day-old chicks trained in a weak passive avoidance learning paradigm

Glucocorticoids are released during learning situations and can trigger neural actions through binding to receptors in different brain areas. The possible role of a glucocorticoid action in long-term memory formation was studied, in day-old chicks, by using a passive avoidance task which chicks otherwise only retain for a few hours (< 10) after training. Thus, we examined the effects of intracerebral corticosterone administration on retention 24 h posttraining. The results showed that chicks injected with corticosterone (1 microgram) at either 15 min pretraining or at 5, 30, 60 min (but not 120, 180, or 360 min) posttraining retained the passive avoidance response when tested 24 h posttraining. Studies with specific mineralocorticoid or glucocorticoid receptor antagonists (RU 28318 or RU 38486, respectively) indicated that this increase in retention by corticosterone might be mediated through glucocorticoid receptors. In order to assess whether the facilitatory effect of corticosterone was mediated through an effect on protein synthesis mechanisms, the protein synthesis inhibitor anisomycin was administered prior to corticosterone. However, this treatment only partially attenuated the effect of the steroid, suggesting that corticosterone may influence other cellular processes involved in the formation of long-term memory for the avoidance behaviour.

A role for the neural cell adhesion molecule in a late, consolidating phase of glycoprotein synthesis six hours following passive avoidance training of the young chick

We have investigated the effect of intracranial injections of the amnestic anti-metabolite, 2-deoxygalactose, and antibodies to the neural cell adhesion molecule on retention of a one-trial passive avoidance task in chicks. Groups of chicks received bilateral intracranial injections of 10 mumol/hemisphere 2-deoxygalactose or 10 microliters/hemisphere anti-neural cell adhesion molecule and were tested 24 h following training. 2-Deoxygalactose injections were amnestic when administered at a previously established time (30 min pre-training). Here we show that the agent is also amnestic when injected within a second time window occurring specifically 6-8 h after training. Administration of 2-deoxygalactose between 2 and 6 h or after 8 h post-training was without effect on retention tested 24 h following training. Anti-neural cell adhesion molecule injections were amnestic only when performed at a time which coincided with the second phase of 2-deoxygalactose susceptibility. Further experiments demonstrated that the neural cell adhesion molecule is one of the molecules into which 2-deoxygalactose is incorporated. Additionally, we investigated the extent of diffusion of 2-deoxygalactose and anti-neural cell adhesion molecule following their injection, with respect to their residence in forebrain loci known to be involved in the memory for passive avoidance. We interpret these data as indicating that two waves of glycoprotein synthesis are necessary for the establishment of long-term memory for the experience of passive avoidance training. The evidence is discussed in the context of earlier results indicating that the two waves involve different glycoprotein species and, possibly, different forebrain regions. We speculate that the late phase of glycoprotein synthesis coincides with, and is required for, modulation of cell-cell adhesion processes, reflecting the selection and stabilization of synapses which maintain an enduring representation of long-term memory.

Reminder effects - reconsolidation or retrieval deficit? Pharmacological dissection with protein synthesis inhibitors following reminder for a passive-avoidance task in young chicks

It is generally accepted that memory formation involves an irreversible passage via labile phases to the stable form of ‘long‐term memory’ impervious to amnestic agents such as protein synthesis inhibitors. However, recent experiments demonstrate that reactivation of memory by way of a reminder renders it labile to such inhibitors, suggesting that such retrieval is followed by a so‐called reconsolidation process similar or identical in its cellular and molecular correlates to that occurring during the initial consolidation. We compared the effects of the protein synthesis inhibitor anisomycin and the glycoprotein synthesis inhibitor 2‐deoxygalactose on the temporal dynamics and pharmacological sensitivity of initial consolidation and memory expression following a reminder in a one‐trial passive‐avoidance task in day‐old chicks. This comparison revealed three differences between the action of the inhibitors on newly formed compared with reactivated memory. First, the recall deficit after the reminder was temporary, whilst the amnesia following inhibitor treatment during training was stable. Second, the sensitive period for the effect of anisomycin was shorter in the reminder than in the training situation. Third, the effective dose for either inhibitor for reminder‐associated amnesia was several times lower than for amnesia developing after training. Thus though like initial consolidation, memory expression at delayed periods following reminder depends on protein and glycoprotein synthesis, the differences between the temporal and pharmacological dynamics in the two situations point to the distinct character of the molecular processes involved in postreminder effects.

An inhibitor of nitric oxide synthesis prevents memory formation in the chick

Memory formation is presumed to require retrograde communication across synaptic junctions. Nitric oxide (NO) is a putative retrograde messenger at N-methyl-D-aspartate (NMDA)-mediated synapses [8, 9]. Inhibitors of nitric oxide synthesis block initiation of long-term potentiation [2, 3, 19]. Memory for a one-trial passive avoidance task in the young chick involves an NMDA-linked intracellular cascade culminating in lasting modulation of synaptic morphology and [6, 18]. Here we show that injection of the nitric oxide synthase inhibitor N-nitro-L-arginine prior to training results in amnesia for the passive avoidance task; the amnesia can be overcome by injecting L-arginine along with the inhibitor. Thus we have verified for the first time experimentally that NO plays a role in memory formation.

Passive avoidance training results in lasting changes in deoxyglucose metabolism in left hemisphere regions of chick brain.

Day-old chicks peck when offered a bright bead; if the bead is coated with the bitter-tasting methylanthranilate (M) they avoid it thereafter. 2-[14C] Deoxyglucose injected 1 min prior to training shows increased uptake into the hyperstriatum ventrale (HV) and lobus parolfactorius (LPO) 30 min later compared with control birds which have pecked a water-coated bead (W). To distinguish effects of training from those of consolidation, and to study lateralization of the increased uptake, 2-[14C]deoxyglucose (4 muCi) was injected ip either 5 min before, or 10 or 30 min after training. Thirty minutes after injection, bilateral samples of medial hyperstriatum ventrale (MHV), LPO and palaeostriatum augmentatum (PA)-enriched regions were dissected. Specific radioactivity (dmp/mg X prot) in left and right MHV and left and right LPO was standardized on the mean PA-specific radioactivity for each bird. When 2-DG was injected 5 min prior to training, standardized radioactivity in the left LPO was 26% greater, and in the left MHV 13% greater in M than W birds. There were no differences in the right hemisphere. With injection 10 min after training, there was an increase of 22% in the left LPO of M birds over W, of 29% in the left MHV and 22% in the right MHV. If injection was delayed to 30 min after training, there was no increase in the LPO, but a 13% increase persisted in the left MHV. Enhanced 2DG metabolism following passive avoidance training is thus persistent, lateralized, and, in the MHV at least may represent an aspect of cellular reorganization consequent on experience but independent of the immediate concomitants of training--perhaps part of the process of memory consolidation.

Learning induced increase of immediate early gene messenger rna in the chick forebrain

The immediate early genes c‐fos and c‐jun are activated rapidly in nerve cells in response to in vivo and in vitro stimulation. Because of their involvement in transcriptional regulation, the products of these genes have been proposed as nuclear signals for consolidation of long‐term memory. However, no specific changes of immediate early gene expression in relation to learning have yet been reported. Compared with quiet controls, training young chicks to discriminate food grains from inedible pebbles results in a 4.8‐fold elevation of c‐fos and a 3.7‐fold elevation of c‐jun mRNA in the forebrain 30 min after an 8‐min training session. Compared to chicks that had learned the discrimination and were merely repeating already learned behaviour, the increase in c‐jun mRNA in the learning group was 64% (P < 0.03) but a 24% increase in c‐fos mRNA was not significant. Because the increased expression is higher in birds that are learning the task than in those that are repeating already learned behaviour, and is not proportional to behavioural activity per se, we conclude that learning a new task is itself responsible for enhanced expression of the genes.

Training induced dendritic spine density changes are specifically related to memory formation processes in the chick, Gallus domesticus

The density of dendritic spines on large, multipolar, projection neurons in the intermediate medial hyperstriatum ventrale (IMHV) of 1-day-old chicks was examined after training on a one-trial passive avoidance task. Chicks trained on the task were given a brief, subconvulsive transcranial electroshock 5 min after training, a procedure which rendered about half of them amnesic. The spine density was found to be significantly higher 24-26 h after training in the left IMHV in chicks that remembered the response compared to chicks rendered amnesic. These data, taken together with our earlier observations on spine density changes following training, argue strongly in favour of a specific role for dendritic spines in memory formation in the chick.

MEMORY FORMATION IN THE CHICK DEPENDS ON MEMBRANE-BOUND PROTEIN KINASE C

The role of protein kinase C (PKC) in the formation of memory for a one-trial passive avoidance task in 1-day-old chicks has been studied, following earlier observations that training on this task results in transient and lateralised changes in the phosphorylation state of presynaptic B-50 protein, a PKC substrate. In accord with hypotheses that the activity of PKC is regulated by translocation from cytosol to membrane, a significant increase was found in the fraction of the alpha/beta forms of the enzyme, assayed immunologically, present in a synaptic-membrane-bound, Triton-extractable form in the left intermediate medial hyperstriatum ventrale (IMHV) of chicks 30 min after training on the passive avoidance task. Two inhibitors of PKC, melittin (10 microliters, 120 microM) and H7 (10 microliters, 10 mM), if injected intracerebrally 10 min prior to or 10 min after training, were without effect on the general behaviour of the chicks or their training. However, these injections of the inhibitors produced amnesia in birds tested 3 h later. This effect was lateralised; only left hemisphere injections of the inhibitors produced amnesia. A possible state-dependency interpretation of these results was ruled out. The results are discussed in the context of hypotheses as to the regulatory role of PKC in neural plasticity and memory formation.

Glycoproteins and memory formation

One-trial passive avoidance training in day-old chicks results in a biochemical cascade occurring in two forebrain regions, the intermediate medial hyperstriatum ventrale and the lobus parolfactorius. This cascade, initiated by synaptic transients, results in the activation of immediate early genes and culminates in the de novo synthesis of a family of pre- and post-synaptic membrane glycoproteins, that, inserted into the membrane, serve in the remodelling of synaptic connectivity which is a requirement for the brain representations constituting long-term memory. There are two waves of glycoprotein synthesis consequent on training, the first occurring within an hour of the training experience and the second 5.5-8 h post-training. Blocking synthesis during these time windows results in amnesia for the task. Amongst the glycoproteins involved are two cell adhesion molecules, NCAM and L1. Injection of antibodies to L1 result in amnesia if injected during either time window, but not outside these times; antibodies to NCAM result in amnesia only if injected at the 5.5-h timepoint. I interpret these results as indicating that de novo synthesis of NCAM during the second time window is necessary for producing a persistent memory trace.