Herman T. Epstein

Bryostatin Enhancement of Memory in Hermissenda

Bryostatin, a potent agonist of protein kinase C (PKC), when administered to Hermissenda was found to affect acquisition of an associative learning paradigm. Low bryostatin concentrations (0.1 to 0.5 ng/ml) enhanced memory acquisition, while concentrations higher than 1.0 ng/ml down-regulated the pathway and no recall of the associative training was exhibited. The extent of enhancement depended upon the conditioning regime used and the memory stage normally fostered by that regime. The effects of two training events (TEs) with paired conditioned and unconditioned stimuli, which standardly evoked only short-term memory (STM) lasting 7 min, were—when bryostatin was added concurrently—enhanced to a long-term memory (LTM) that lasted about 20 h. The effects of both 4- and 6-paired TEs (which by themselves did not generate LTM), were also enhanced by bryostatin to induce a consolidated memory (CM) that lasted at least 5 days. The standard positive 9-TE regime typically produced a CM lasting at least 6 days. Low concentrations of bryostatin (<0.5 ng/ml) elicited no demonstrable enhancement of CM from 9-TEs. However, animals exposed to bryostatin concentrations higher than 1.0 ng/ml exhibited no behavioral learning. Sharp-electrode intracellular recordings of type-B photoreceptors in the eyes from animals conditioned in vivo with bryostatin revealed changes in input resistance and an enhanced long-lasting depolarization (LLD) in response to light. Likewise, quantitative immunocytochemical measurements using an antibody specific for the PKC-activated Ca2+/GTP-binding protein calexcitin showed enhanced antibody labeling with bryostatin. Animals exposed to the PKC inhibitor bisindolylmaleimide-XI (Ro-32-0432) administered by immersion prior to 9-TE conditioning showed no training-induced changes with or without bryostatin exposure. However, if animals received bryostatin before Ro-32, the enhanced acquisition and demonstrated recall still occurred. Therefore, pathways responsible for the enhancement effects induced by bryostatin were putatively mediated by PKC. Overall, the data indicated that PKC activation occurred and calexcitin levels were raised during the acquisition phases of associative conditioning and memory initiation, and subsequently returned to baseline levels within 24 and 48 h, respectively. Therefore, the protracted recall measured by the testing regime used was probably due to bryostatin-induced changes during the acquisition and facilitated storage of memory, and not necessarily to enhanced recall of the stored memory when tested many days after training.

Time windows for effects of protein synthesis inhibitors on Pavlovian conditioning in Hermissenda: Behavioral aspects

Inhibitors of protein and mRNA syntheses inhibit long-term memory (LTM), but we lack information about the time windows during which those inhibitors are effective. Anisomycin (a protein synthesis translation inhibitor) was given to Hermissenda crassicornis which had received sufficient Pavlovian conditioning to produce LTM. When tested after 90 min, LTM recall was blocked when anisomycin was administered until 13 min after conditioning and from 65 to 75 min. There was good recall from 16 to 60 min. When tested at 240 min, two periods of sensitivity to anisomycin were revealed: one finished before 20 min, while the other lasted from 60 to 220 min.A brief study of the transcription inhibitor actinomycin-D found its action similar to that of anisomycin, except not inhibiting recall from about 160 to 220 min.

Stages of increased cerebral blood flow accompany stages of rapid brain growth

Stages of increased cerebral blood flow accompany stages of rapid brain growth The existence of stages of rapid brain growth implies the existence of associated stages of increased cerebral blood flow (CBF) to supply the substances and energy needed for the added brain weight. Studies in the literature give developmental data for CBF in humans which show stages of increased cerebral blood flow at ages starting just preceding and/or coincident with the ages of the rapid brain growth stages. Confirmation of the implication for one of the earliest stages is also found in PET data. Additionally, data for rodents show a similar association of stages of increasing CBF and their brain growth stages. Thus, to the accuracy of the data, the implication is confirmed. Finding stages in cerebral blood flow predicts the existence of stages of brain growth. In addition; finding correlated blood flow stages also supports the age spans of the stages of rapid brain growth in both species. Such correlations also suggest that measurements of blood flow anywhere in the body might serve to reveal rapid growth stages in particular localities or organs if the blood flow shows significant stages. The ratio of energy consumed in brain to that in total body decreases from close to 100% at birth to the adult value of about 20% by age 18 years.

An Outline of the Role of Brain in Human Cognitive Development

Brain development in humans occurs stagewise in correlation with the onsets of the main Piagetian stages of reasoning development. This allows a description of cognitive development as at least partially resulting from and dependent upon biological events occurring in the brain. Evidence shows that some eventual brain structures depend on a combination of biological events and instructional inputs. This contributes to the instruction dependence of the higher cognitive functions. Such a description thereby permits some novel working hypotheses about normal cognitive development and how to foster it as well as suggests alternative ways of creating intervention programs for children from deprived environments.

Pavlovian conditioning-specific increases of the Ca2+- and GTP-binding protein, calexcitin in identified Hermissenda visual cells.

Hermissenda CNS, immunolabeled for the memory protein calexcitin showed significant immunostaining over background in the B-photoreceptor cells of the eye. The degree of staining correlated positively with the number of Pavlovian training events experienced by the animals and the degree of Pavlovian conditioning induced. The training regime consisted of exposing animals to light (conditioned stimulus, CS) paired with orbital rotation (unconditioned stimulus, US). In animals that exhibited the conditioned response, calexcitin immunolabeling was more intense than was found for naive (unconditioned) animals or animals given the CS and US in random sequence. Animals exposed to lead (maintained in 1.2 ppm lead acetate) at a dosage known to impair learning in children, showed reduced learning and less intense calexcitin staining whether the CS and US were paired or given randomly. However, the levels were still higher than that of naive animals. Immuno-electron microscopy indicated that the labeling was predominantly within calcium sequestering organelles such as the endoplasmic reticulum, and to lesser extent within mitochondria, and photopigments. The calexcitin density after a short-term memory (STM) regime was the same whether measured 5 minutes after conditioning (when STM was evidenced by foot contraction) or 90 minutes later when no recall was detected. The staining density was also similar to the levels found 5 minutes after long-term memory (LTM) conditioning. However, the LTM regime produced a greater calexcitin intensity at 90 minutes when the memory had been consolidated. This learning-specific increase in calexcitin is consistent with the previously implicated sequence of molecular events that are associated with progressively longer time domains of memory storage.

Two different biological configurations for long-term memory

Long-term memory (LTM) in Hermissenda can be distinguished from consolidated long-term memory (CLTM) by determining how long recall is retained. LTM is retained for approximately 1 day, while CLTM is retained for at least 3 days. During the transition from LTM to CLTM, the extent of retention appears to depend partially on how much consolidation has been completed. Several models are discussed that may be related to the two different manifestations of recall.

Lead, Learning, and Calexcitin in Hermissenda

widely different; i.e., a Ki should be determined for Leu-Pro. The third conclusion suggests that a new criterion for the socalled squid type OPAA has emerged, namely, the lack of inhibition by Leu-Pro. Two questions are now to be examined: Do either Leu or Pro, as hydrolysis products, inhibit the Mazur type OPAAs? And if so, do any other amino acids inhibit the squid type OPAA? Because the latter category now includes the enzyme ird, for the first time, a bacteri l OPAA, namely that from seudomonas dim nuta, shows a close resemblance to the squid zyme. The first two conclusions, while appearing valid, from Pseudomonas diminuta as well as from squid nerve, a new approach to the question of a natural substrate for this enzyme may be emerging.

Evolution of the reasoning hominid brain

Cognition is readily seen to be connected to evolution through plots of the ratio of cranial capacity to body size of hominids which show two regions of sharply increasing ratios beginning at 2.5 and 0.5 million years ago – precisely the critical times inferred by the author from his study of tools. A similar correlation exists between current human brain growth spurts and the onsets of the Piagetian stages of reasoning development. The first goal of the authors target article is stated to be “to make a case for the relevance of archeological contributions to studies of the evolution of cognition” (sect. 1, Introduction). His analysis focuses on spatial cognition.

Training Alone, Not the Tripeptide RGD, Modulates Calexcitin in Hermissenda

In the nudibranch mollusc Hermissenda crassicornis, the intensity of immunostaining for the Ca2+/GTP-binding protein calexcitin (1) correlates positively with the degree of learning obtained and the level of memory expressed after Pavlovian conditioning (2, 3). When inhibitors of transcription (actinomycin-D; Act-D) and translation (anisomycin; ANI) are applied after training, they affect the animals ability to recall the learned behavior. Epstein et al. (4) have recently described the time windows for these effects. They noted that there are two phases of sensitivity to the inhibitors: an early phase, immediately after training (0-13 min), and a later phase (70-160 min for Act-D, and 70-220 min for ANI). Subsequently, Epstein et al. (5) reported that long-term memory also has two distinct biological configurations: long-term memory (LTM), which lasts about 24 h, and consolidated long-term memory (CLTM), which persists up to 6 days. The amount of memory retained by conditioned animals appears to depend, in part, on how much consolidation has been completed; and the transition from LTM to CLTM is sensitive to a cell adhesion molecule (CAM) inhibitor, the tripeptide arginyl-glycyl-aspartate (RGD). One working hypothesis predicts that increased calexcitin levels are needed to establish LTM, and that RGD-sensitive CAMs are subsequently involved in the transition from LTM to CLTM. What was unknown was the possible relationship between calexcitin levels and the effects of RGD during this critical transition period. Thus, as part of a continuing study to describe and define the memory stages demonstrated in Hermissenda (6, 4, 5), we undertook an immunocytochemical study to investigate possible correlations between calexcitin levels and the observed effects of RGD

Lead affects learning by Hermissenda crassicornis

Figure 1. (A) Temporal transfer functions measured by Batra and Barlow (dashed line), Brodie et al. (solid line), and in our preliminary experiments with filled circles) and without (unfilled circles) background illumination. (B) TTFfrom three single ommatidia in as many eyes. Filled circles denote responses recorded with constant background illumination and unfilled circles those recorded without background. Error bars denote standard deviation.