A feedback-controlled ensemble model of the stress-responsive hypothalamo-pituitary-adrenal axis

The present work develops and implements a biomathematical statement of how reciprocal connectivity drives stress-adaptive homeostasis in the corticotropic (hypothalamo-pituitary-adrenal) axis. In initial analyses with this interactive construct, we test six specific a priori hypotheses of mechanisms linking circadian (24-h) rhythmicity to pulsatile secretory output. This formulation offers a dynamic framework for later statistical estimation of unobserved in vivo neurohormone secretion and within-axis, dose-responsive interfaces in health and disease. Explication of the core dynamics of the stress-responsive corticotropic axis based on secure physiological precepts should help to unveil new biomedical hypotheses of stressor-specific system failure.

Ensemble codes involving hippocampal neurons are at risk during delayed performance tests

Multielectrode recording techniques were used to record ensemble activity from 10 to 16 simultaneously active CA1 and CA3 neurons in the rat hippocampus during performance of a spatial delayed-nonmatch-to-sample task. Extracted sources of variance were used to assess the nature of two different types of errors that accounted for 30% of total trials. The two types of errors included ensemble “miscodes” of sample phase information and errors associated with delay-dependent corruption or disappearance of sample information at the time of the nonmatch response. Statistical assessment of trial sequences and associated “strength” of hippocampal ensemble codes revealed that miscoded error trials always followed delay-dependent error trials in which encoding was “weak,” indicating that the two types of errors were “linked.” It was determined that the occurrence of weakly encoded, delay-dependent error trials initiated an ensemble encoding “strategy” that increased the chances of being correct on the next trial and avoided the occurrence of further delay-dependent errors. Unexpectedly, the strategy involved “strongly” encoding response position information from the prior (delay-dependent) error trial and carrying it forward to the sample phase of the next trial. This produced a miscode type error on trials in which the “carried over” information obliterated encoding of the sample phase response on the next trial. Application of this strategy, irrespective of outcome, was sufficient to reorient the animal to the proper between trial sequence of response contingencies (nonmatch-to-sample) and boost performance to 73% correct on subsequent trials. The capacity for ensemble analyses of strength of information encoding combined with statistical assessment of trial sequences therefore provided unique insight into the “dynamic” nature of the role hippocampus plays in delay type memory tasks.

Random circular permutation of genes and expressed polypeptide chains: application of the method to the catalytic chains of aspartate transcarbamoylase.

Recent studies on proteins whose N and C termini are in close proximity have demonstrated that folding of polypeptide chains and assembly of oligomers can be accomplished with circularly permuted chains. As yet no methodical study has been conducted to determine how extensively new termini can be introduced and where such termini cannot be tolerated. We have devised a procedure to generate random circular permutations of the catalytic chains of Escherichia coli aspartate transcarbamoylase (ATCase; EC 2.1.3.2) and to select clones that produce active or stable holoenzyme containing permuted chains. A tandem gene construct was made, based on the desired linkage between amino acid residues in the C- and N-terminal regions of the polypeptide chain, and this DNA was treated with a suitable restriction enzyme to yield a fragment containing the rearranged coding sequence for the chain. Circularization achieved with DNA ligase, followed by linearization at random with DNase I, and incorporation of the linearized, repaired, blunt-ended, rearranged genes into a suitable plasmid permitted the expression of randomly permuted polypeptide chains. The plasmid with appropriate stop codons also contained pyrI, the gene encoding the regulatory chain of ATCase. Colonies expressing detectable amounts of ATCase-like molecules containing permuted catalytic chains were identified by an immunoblot technique or by their ability to grow in the absence of pyrimidines in the growth medium. Sequencing of positive clones revealed a variety of novel circular permutations. Some had N and C termini within helices of the wild-type enzyme as well as deletions and insertions. Permutations were concentrated in the C-terminal domain and only few were detected in the N-terminal domain. The technique, which is adaptable generally to proteins whose N and C termini are near each other, can be of value in relating in vivo folding of nascent, growing polypeptide chains to in vitro renaturation of complete chains and determining the role of protein sequence in folding kinetics.