Pharmaco-metabonomic phenotyping and personalized drug treatment

There is a clear case for drug treatments to be selected according to the characteristics of an individual patient, in order to improve efficacy and reduce the number and severity of adverse drug reactions. However, such personalization of drug treatments requires the ability to predict how different individuals will respond to a particular drug/dose combination. After initial optimism, there is increasing recognition of the limitations of the pharmacogenomic approach, which does not take account of important environmental influences on drug absorption, distribution, metabolism and excretion. For instance, a major factor underlying inter-individual variation in drug effects is variation in metabolic phenotype, which is influenced not only by genotype but also by environmental factors such as nutritional status, the gut microbiota, age, disease and the co- or pre-administration of other drugs. Thus, although genetic variation is clearly important, it seems unlikely that personalized drug therapy will be enabled for a wide range of major diseases using genomic knowledge alone. Here we describe an alternative and conceptually new 'pharmaco-metabonomic' approach to personalizing drug treatment, which uses a combination of pre-dose metabolite profiling and chemometrics to model and predict the responses of individual subjects. We provide proof-of-principle for this new approach, which is sensitive to both genetic and environmental influences, with a study of paracetamol (acetaminophen) administered to rats. We show pre-dose prediction of an aspect of the urinary drug metabolite profile and an association between pre-dose urinary composition and the extent of liver damage sustained after paracetamol administration.     

Symmetrical directional cloning: An efficient method to prepare hairpin RNA interference constructs

RNA interference (RNAi) is a loss-of-function approach by which double-stranded RNA (dsRNA) initiates degradation of homologous mRNAs in a sequence specific manner. The dsRNA molecules can be produced in vitro or in vivo, and can be introduced to cells in a number of ways. Here we report a more efficient method for the cloning of inverted repeat DNA fragments into expression vectors that can be transcribed into effective dsRNA molecules in vivo or in vitro. This method, named Symmetrical Directional Cloning (SDC), takes the advantage of compatible non-palindromic restriction enezyme sites, which allow one to directionally clone a single PCR product in both the sense and antisense orientations together into a vector. SDC allows for the directional cloning of inverted repeats using a single PCR product; it requires only one cut site on each side of the loop. Hence this method is more cost effective and less time-consuming. At least 21 commercially available restriction endonucleases can be used as cloning sites for the SDC method. The efficacy of dsRNA expression vectors prepared by SDC has been demonstrated by targeting a negative regulator of the signaling pathway mediating the response of cells to phytohormone, gibberellins (GA), in the aleurone cells.     

A presynaptic action of glutamate at the cone output synapse

Neurotransmitter release from many central nervous system synapses is regulated by 'autoreceptors' at the synaptic terminal, which bind the released transmitter and alter release accordingly. The photoreceptors of lower vertebrates are thought to use glutamate as a neurotransmitter. Glutamate conveys the visual signal to postsynaptic bipolar and horizontal cells, but has been reported not to act on the photoreceptors themselves. We show here that glutamate evokes a current, carried largely by chloride ions, in cones isolated from the tiger salamander retina. This response is localized to the synaptic terminal of the cone. Removing external sodium blocks this action of glutamate. These results suggest the existence of a positive feedback loop at the cone output synapse: over most of the light-response range, glutamate released by depolarization of the cone will cause further depolarization, increasing the gain of phototransduction. Glutamate released from rods may also polarize cones, modulating the gain of the cone output synapse. This system is surprisingly different from the autoreceptor systems for most other transmitters, which act in a negative feedback way.     

Understory vegetation in fully stocked pinyon-juniper stands

Ten fully stocked pinyon-juniper stands contained a total of 73 species in the understory, but the number of understory species in any one stand was moderately low (mean = 20). On each stand, species of at least five different plant groups were present in the understory (shrub, perennial grass, perennial forb, annual grass, or annual forb). A perennial grass, Sandberg bluegrass ( Poa sandbergii ), and a group of annual forbs with relatively high cover and constancy among stands appeared best adapted to coexist with the pinyon-juniper overstory. The proportion of total plant cover was greater on tree-associated microsites (duff and transition) than in the interspace between trees because of the greater surface area of the former in most stands. The transition microsite was the most favorable for understory species and provided understory cover in disproportionately greater amounts than the area it occupied.     

Understory seed rain in harvested pinyon-juniper woodlands

Seed rain was collected on six paired tree harvest and undisturbed plots in singleleaf pinyon ( Pinus monophylla )–Utah juniper ( Juniperus osteosperma ) stands. Approximately 14,600 seeds were collected during four years. Seed rain in undisturbed plots was similar to levels in mixed forest communities. Seed rain on harvest plots was similar to disturbed sites and grasslands. Seed rain levels reflect the current successional stage rather than the climax community type for the site. Seed rain increased in numbers and seed production per unit of plant cover following tree removal and especially on transition soil microsites. Only three to four of the plant species present on a site contributed greater than 10% of the total seed rain. Seed rain composition was similar on harvest and undisturbed plots (Jaccard Similarity Index Values = 47% to 67%) and explains in part the rapid reestablishment of predisturbance understory communities.     

Understory response to tree harvesting of singleleaf pinyon and Utah juniper

Fifteen widely separated sites within the pinyon - juniper woodlands of the Great Basin were cleared of trees. Understory response was recorded for 2 to 4 years. The array of postharvest plant assemblages were classified into one of four phytosociological groups using discriminant analysis. Pre - and postharvest plant assemblages from the same site appeared in the same phytosociological group, which indicates postharvest response could be predicted from the preharvest plant assemblage. Initial postharvest response appears cyclic in nature, and cycles are controlled by both residual plants and the rapid immigration of shrub species. Perennial plant density generally declined following tree harvest, but the fewer remaining plants produced significantly more cover than in preharvest stands.         

Changes in brain dopamine levels and aggressive behavior with aging in 2 mouse strains

The genetic programming of brain monoamine changes with aging show remarkable differences in 2 mouse strains. A marked increase in dopamine occurred in 32-week-old grouped ICR mice and the males showed intense irritability and aggressive behavior. Brain amines changed only slightly in old C57BL6J mice and behavior remained benign. Old females showed similar amine changes but aggresive behavior did not occur in either strain.     

Changes in brain dopamine levels and aggressive behavior with aging in 2 mouse strains

Summary The genetic programming of brain monoamine changes with aging show remarkable differences in 2 mouse strains. A marked increase in dopamine occurred in 32-week-old grouped ICR mice and the males showed intense irritability and aggressive behavior. Brain amines changed only slightly in old C57BL6J mice and behavior remained benign. Old females showed similar amine changes but aggressive behavior did not occur in either strain.