Noncontact dipole effects on channel permeation. VI. 5F- and 6F-Trp gramicidin channel currents

Fluorination of peptide side chains has been shown to perturb gramicidin channel conductance without significantly changing the average side chain structure, which, it is hoped, will allow detailed analysis of electrostatic modulation of current flow. Here we report a 1312-point potassium current-voltage-concentration data set for homodimeric channels formed from gramicidin A (gA) or any of eight fluorinated Trp analogs in both lecithin and monoglyceride bilayers. We fit the data with a three-barrier, two-site, two-ion (3B2S) kinetic model. The fluorination-induced changes in the rate constants were constrained by the same factor in both lipids. The rate constant changes were converted to transition-state free-energy differences for comparison with previous electrostatic potential energy differences based on an ab initio force field. The model allowed a reasonably good fit (chi = 8.29 with 1271 degrees of freedom). The measured changes were subtle. Nevertheless, the fitted energy perturbations agree well with electrostatic predictions for five of the eight peptides. For the other three analogs, the fitted changes suggested a reduced translocation barrier rather than the reduced exit barrier as predicted by electrostatics.     

A squirrel monkey model of poststroke motor recovery

Nonhuman primate models of poststroke recovery have become increasingly rare primarily due to high purchase and maintenance costs and limited availability of nonhuman primate species. Despite this obstacle, nonhuman primate models may offer important advantages over rodent models for understanding many of the brain's mechanisms for self-repair due to greater similarity in cortical organization to humans. Since the mid-1990s, surgical, neurophysiological, and neuroanatomical methods have been developed to understand structural and functional remodeling of the cerebral cortex after an ischemic event, such as occurs in stroke. These methods require long surgical procedures and entail constant physiological monitoring. With careful attention to intraoperative and postsurgical monitoring, these procedures can be repeated multiple times in individual monkeys without untoward events. This model provides a statistically powerful approach for tracking brain plasticity in the ensuing weeks and months after a stroke-like injury, reducing the number of animals required for individual experiments. This methodology is described in detail, and many of the resulting findings that are relevant for understanding stroke recovery and the effects of rehabilitative and pharmacotherapeutic interventions are summarized.     

A cellular mechanism for prepulse inhibition

In prepulse inhibition (PPI), startle responses to sudden, unexpected stimuli are markedly attenuated if immediately preceded by a weak stimulus of almost any modality. This experimental paradigm exposes a potent inhibitory process, present in nervous systems from invertebrates to humans, that is widely considered to play an important role in reducing distraction during the processing of sensory input. The neural mechanisms mediating PPI are of considerable interest given evidence linking PPI deficits with some of the cognitive disorders of schizophrenia. Here, in the marine mollusk Tritonia diomedea, we describe a detailed cellular mechanism for PPI--a combination of presynaptic inhibition of startle afferent neurons together with distributed postsynaptic inhibition of several downstream interneuronal sites in the startle circuit.     

Growth hormone and osteoporosis: an overview of endocrinological and pharmacological insights from the Utah paradigm of skeletal physiology

Multidisciplinary advances in skeletal physiology offer a new paradigm for the effects of growth hormone (GH) and other agents on bone and osteoporosis. The still-evolving Utah paradigm of skeletal physiology supplements earlier ideas with later discovered roles of the skeleton's tissue-level 'nephron equivalents' and muscle strength in skeletal development, physiology and disorders. This article summarizes how these factors could influence the effects of GH on bone strength and bone 'mass', and the use of GH in the treatment of osteoporoses. Although the cellular and molecular biological mechanisms involved remain obscure, the associated cascades of cellular, genetic and biochemical processes and molecules should offer many opportunities to find or design agents that have medically useful effects on bone and muscle without giving rise to unwanted side-effects.     

Why the ISMNI and the Utah paradigm? Their role in skeletal and extraskeletal disorders

Besides bringing problems, aging can let the mind's eye see more clearly than before, and it can let us express ourselves better. As age, experience and common sense examine today's skeletal medicine and surgery two questions keep popping up: A) How did we fail?; B) How to make it better? The Utah paradigm of skeletal physiology and the seminal ISMNI offer some answers, but exploiting them faces problems. Problem #1: By 1960 all clinicians and physiologists 'knew' (as the ancients 'knew' this world is flat) that effector cells controlled solely by nonmechanical agents explain all skeletal physiology and disorders ('effector cells' include osteoblasts, osteoclasts, chondroblasts and fibro-blasts). Or, nonmechanical agents -->cell level -->organ and intact subject. Adding later-discovered information to that 1960 view led to the Utah paradigm, which reveals the formerly hidden tissue-level 'dimension' of skeletal physiology. It builds on this idea: (mechanical + nonmechanical agents) -->(tissue level + cell level) --> organ and intact subject. The paradigm assigns great influence of neuromuscular physiology and physical activities on skeletal architecture, strength and mechanical competence. It also exposes flaws in many older views so controversies arise. Problem #2: The Utah paradigm and Wegner's concept of plate tectonics in geology seem alike in that each is valid but came before its time, so others fought it. They differ in this: The fight about Wegner's idea is over, but for the Utah paradigm and the ISMNI it just began. Hence more controversies. Nevertheless: A growing minority realizes that paradigm provides a far better base to build on than its antecedents, and since it keeps evolving as more evidence comes in it could endure for some decades. Yet very few realize this: It and the ISMNI have important implications for fields besides biomechanics and orthopaedics. Examples include anatomy, cardiovascular disease, dentistry, endocrinology, family medicine, gastroenterology, general surgery, genetics, gerontology, gynecology, maxillofacial surgery, neurology, neurosurgery, nutrition, ophthalmology, pathology, pediatrics, physical medicine and rehabilitation, plastic surgery, radiology, rheumatology, space and sports medicine, and urology. Quite a list! For the italicized questions above this article offers answers, of which its conclusion distills an essence.     

Mucosal challenge of Macaca nemestrina with simian immunodeficiency virus (SIV) following SIV nucleocapsid mutant DNA vaccination

A simian immunodeficiency virus (SIV)(Mne) DNA clone was constructed that produces viruses containing a four amino acid deletion in the second zinc finger of the nucleocapsid (NC) domain of the Gag polyprotein. Viruses produced from this clone, although non-infectious both in vitro and in vivo, complete a majority of the steps in a single retroviral infection cycle. Eight pig-tailed macaques (Macaca nemestrina) were inoculated intramuscularly and subcutaneously three times over the course of 24 weeks with the NC mutant expressing DNA. These macaques, and four controls, were then challenged mucosally (intrarectally) with the homologous virus (SIV Mne CL E11S) and monitored for evidence of infection and clinical disease. Prior to challenge, a measurable humoral immune response was noted in four of eight immunized macaques. After challenge, all 12 macaques became infected, although four immunized animals greatly restricted their viral replication, and one immunized animal that controlled replication remains antibody negative. No disease has been evidence during the 46-week period of monitoring after challenge.     

Expression of beta-defensin-1 in chimpanzee (Pan troglodytes) airways

In human airways, beta-defensins function in the elimination of various pathogens. They have been identified in a wide range of species. Here we report the identification and expression of chimpanzee beta-defensin-1 (cBD1), which is a homolog of human beta-defensin-1, in chimpanzee airways and skin. The cBD1 cDNA sequence differs by only one synonymous nucleotide substitution compared to the human cDNA sequence. In situ hybridization revealed that in lung tissue beside alveolar macrophages also airway epithelial cells, endothelial cells and type II pneumocytes express cBD1 mRNA. In skin, cBD1 mRNA was expressed in keratinocytes and endothelial cells. Together, these results show similarity in structure and expression pattern and perhaps in function.     

Genetic Analysis of the Role of the Transfer Gene, traN, of the F and R100-1 Plasmids in Mating Pair Stabilization during Conjugation

Mating pair stabilization occurs during conjugative DNA transfer whereby the donor and recipient cells form a tight junction which requires pili as well as TraN and TraG in the donor cell. The role of the outer membrane protein, TraN, during conjugative transfer was examined by introduction of a chloramphenicol resistance cassette into the traN gene on an F plasmid derivative, pOX38, to produce pOX38N1::CAT. pOX38N1::CAT was greatly reduced in its ability to transfer DNA, indicating that TraN plays a greater role in conjugation than previously thought. F and R100-1 traN were capable of complementing pOX38N1::CAT transfer equally well when wild-type recipients were used. F traN, but not R100-1 traN, supported a much lower level of transfer when there was an ompA mutation or lipopolysaccharide (LPS) deficiency in the recipient cell, suggesting receptor specificity. The R100-1 traN gene was sequenced, and the gene product was found to exhibit 82.3% overall similarity with F TraN. The differences were mainly located within a central region of the proteins (amino acids 162 to 333 of F and 162 to 348 of R100-1). Deletion analysis of F traN suggested that this central portion might be responsible for the receptor specificity displayed by TraN. TraN was not responsible for TraT-dependent surface exclusion. Thus, TraN, and not the F pilus, appears to interact with OmpA and LPS moieties during conjugation, resulting in mating pair stabilization, the first step in efficient mobilization of DNA.     

Ephemeral bio-engineers or reef-building polychaetes: how stable are aggregations of the tube worm Lanice conchilega (Pallas, 1766)?

Dense aggregations of tube-worms can stabilize sediments and generate oases for benthic communities that are different and often more diverse and abundant than those of the surroundings. If these features are to qualify as biogenic reefs under nature-conservation legislation such as the EC Habitats Directive, a level of stability and longevity is desirable aside from physical and biological attributes. Lanice conchilega (Pallas, 1766) is widely distributed around the European coast and aggregations of this tube-dwelling polychaete are known to have a positive effect on the biodiversity of associated species in inter- and sub-tidal areas. This increases the value of L. conchilega-rich habitats for higher trophic levels such as birds and fish. However, L. conchilega is currently not recognized as a reef builder primarily due to uncertainty about the stability of their aggregations. We carried out three studies on different spatial and temporal scales to explore a number of properties relating to stability: (1) Individual aggregations of L. conchilega of ～1 m(2) were monitored for up to 1 year, (2) records of L. conchilega from a 258-ha area over a 35-year period were analyzed, (3) the recovery of a population of L. conchilega subjected to disturbances by cultivation of Manila clams (Ruditapes philippinarum) was followed over 3 years. The studies provided evidence about the longevity of L. conchilega aggregations, their resistance to disturbance, their resilience in recovering from negative impact and their large-scale persistence. The results showed that populations of L. conchilega were prone to considerable fluctuation and the stability of aggregations depended on environmental factors and on recruitment. The tube-worms proved to be susceptible to disturbance by cultivation of Manila clams but demonstrated the potential to recover from that impact. The long-term monitoring of a large L. conchilega population in the Bay of Mont Saint Michel (France) indicated that aggregations can persist over many decades with a constant, densely populated core area and an expanding and contracting more thinly populated fringe zone. The stability of aggregations of L. conchilega and the structures they form do not unequivocally fit the currently accepted definition of a reef. However, given L. conchilega's accepted reef-like potential to influence diversity and abundance in benthic communities, we suggest clarifying and expanding the definition of reefs so that species with records of significant persistence in particular areas and which otherwise meet expectations of reefs are included within the definition.