Muscarinic acetylcholine receptor produced in recombinant baculovirus infected Sf9 insect cells couples with endogenous G-proteins to activate ion channels.

Following the infection of insect ovarian cells (Sf9) with recombinant bearing the cDNA coding for the rat muscarinic acetylcholine (ACh) receptor subtype m3, ionic flux across the membrane in response to the application of ACh was examined electrophysiologically. We show that ACh activates potassium currents. The response is abolished when cells are treated with pertussis toxin. No ACh-induced currents are observed from uninfected cells or cells infected with virus which do not contain the cDNA coding for ACh receptors in its genome. The characteristics of single channel currents show time-dependent changes following the application of ACh. Initially, ACh activates brief channel currents with a conductance of about 5 pS. The conductance level of channels gradually increases in steps to 10 pS and then to 20 pS and 40 pS. At the same time, channel open probability also increases. Thereafter, additional channels appear, opening and closing independently of, or at times in synchrony with, the original channel.     

Photoreceptor membrane turnover in the crayfish Cherax destructor: electron microscopy and anti-rhodopsin electron-microscopic immunocytochemistry

Summary Examination of the ultrastructure of retinula cells of the Australian crayfish Cherax destructor at different times over a 24-hour cycle, together with patterns of anti-rhodopsin antigenicity, has lead to the formulation of a model of photoreceptor membrane turnover in these animals. Its main features are: (a) the existence of two bursts of rhabdomeral membrane breakdown; one, light-sensitive and synchronous, occurring at dawn, the other, constituting the first part of the membrane replacement phase itself, occurring during the afternoon and night, (b) the desynchronisation of the replacement phase of turnover between animals and to a lesser extent between cells of the same retina, (c) confinement of ultrastructurally detectable signs of photoreceptor membrane processing to the retinula cells themselves, and (d) replacement of a substantial part if not all of the rhabdomeral membrane daily. This model is compatible with many of the observations reported on the American crayfish Procambarus, and utilises the same basic mechanisms that are believed to operate in photoreceptor membrane turnover in many other arthropod compound eyes.     

Efficacy and deficiencies of rapid biomonitoring in biodiversity conservation: a case study in South Africa

Rapid biomonitoring protocols, using biotic indices based on macroinvertebrate diversity to assess river ecosystem health, are widely used globally. Such quick assessment techniques are lauded for the rapid results obtained and the relatively easy protocol used to achieve an answer. However, do such quick assessments of water quality give enough information about ecosystems? Are important details being overlooked? When should a full faunal survey be used in preference? Important research programmes, including environmental impact studies, often misuse biomonitoring techniques, making influential management decisions using superficial, low-level data obtained using biomonitoring tools, inappropriate to address those management objectives. The value of using biomonitoring as a quick tool, versus a more detailed faunal assessment, is considered here. The assessment of teloganodid mayfly fauna occurring in South African rivers provides an example of the value of detailed studies versus superficial family level investigations, showing that a rapid biomonitoring approach should not be used as a shortcut when a more detailed survey is needed. Each situation should be assessed for its own merit in a given set of project circumstances. A checklist of criteria is presented, giving guidance on when rapid biomonitoring alone is valuable and when more detailed assessments would give a more relevant result.     

Identifying differentially methylated genes using mixed effect and generalized least square models

Background  

DNA methylation plays an important role in the process of tumorigenesis. Identifying differentially methylated genes or CpG islands (CGIs) associated with genes between two tumor subtypes is thus an important biological question. The methylation status of all CGIs in the whole genome can be assayed with differential methylation hybridization (DMH) microarrays. However, patient samples or cell lines are heterogeneous, so their methylation pattern may be very different. In addition, neighboring probes at each CGI are correlated. How these factors affect the analysis of DMH data is unknown.     

Changes in rat adrenal cyclic nucleotides during normal and neoplastic growth

In an attempt to correlate changes in cyclic nucleotide levels with in vivo growth of the rat adrenal gland we have measured adrenal cyclic AMP and cyclic GMP in normal, hyperplastic, and neoplastic rat adrenals. The first group of animals were subject to either unilateral adrenalectomy (ADX) or acute hypophysectomy 1 h prior to unilateral adrenalectomy (HADX). Cyclic nucleotides were measured in the contralateral adrenal post-operatively. In HADX rats cyclic GMP rose steadily throughout the 7 day study period, while ADX rats exhibited significant decreases in adrenal cyclic GMP. Cyclic AMP remained approximately 1.5 pm/mg tissue in HADX rats, while in ADX rats there was significant elevation of adrenal cyclic AMP at all time points. Cyclic GMP/cyclic AMP ratios remained constant in HADX animals; however, the growing adrenals of ADX animals exhibited depressed cyclic GMP/cyclic AMP ratios at all time periods.Adrenal hyperplasia was induced in a seond group of animals by a transplantable, corticotropin-secreting, pituitary tumor. Adrenals from age-matched animals served as controls. Adrenal cyclic AMP was significantly elevated in tumor-bearers at a time correspinding to the peak accumulation of adrenal weight, protein and DNA in these animals. In contrast, adrenal cyclic GMP in both tumor-beares and control animals fell steadily throughout the study period. Cyclic GMP/cyclic AMP ratios of control animals decreased from 2 to 3 weeks post-transplant remaining at the 3 week value during the period corresponding to rapid adrenal growth in tumor-bearers. The cyclic GMP/cyclic AMP ratio in the hyperplastic adrenals of tumor-bearers decreased steadily throughout their rapid growth period, suggesting a positive correlation between adrenal growth and depression of the cyclic GMP/cyclic AMP ratio.Cyclic nucleotide levels in neoplastic adrenals of rats bearing the transplantable adrenocortical carcinoma 494 were compared with cyclic nucleotides in normal rat adrenal glands. Cyclic AMP was not different in the two groups. However, the cyclic GMP content of neoplastic adrenals was significantly lower than that of normal adrenal tissue, causing a suppression of the cyclic GMP/cyclic AMP ratio in the neoplastic tissue. Thus, measurement of adrenal cyclic nucleotides in both hyperplastic and neoplastic rat adrenal glands suggests that adrenal growth in vivo may be characterized by a depression of the cyclic GMP/cyclic AMP ratio.     

Arthropod phylogeny: onychophoran brain organization suggests an archaic relationship with a chelicerate stem lineage

Neuroanatomical studies have demonstrated that the architecture and organization among neuropils are highly conserved within any order of arthropods. The shapes of nerve cells and their neuropilar arrangements provide robust characters for phylogenetic analyses. Such analyses so far have agreed with molecular phylogenies in demonstrating that entomostracans+malacostracans belong to a clade (Tetraconata) that includes the hexapods. However, relationships among what are considered to be paraphyletic groups or among the stem arthropods have not yet been satisfactorily resolved. The present parsimony analyses of independent neuroarchitectural characters from 27 arthropods and lobopods demonstrate relationships that are congruent with phylogenies derived from molecular studies, except for the status of the Onychophora. The present account describes the brain of the onychophoran Euperipatoides rowelli, demonstrating that the structure and arrangements of its neurons, cerebral neuropils and sensory centres are distinct from arrangements in the brains of mandibulates. Neuroanatomical evidence suggests that the organization of the onychophoran brain is similar to that of the brains of chelicerates.     

Ischemia-reperfusion injury changes the dynamics of Ca2+-contraction coupling due to inotropic drugs in isolated hearts.

Positive inotropic drugs may attenuate or exacerbate the deleterious effects of ischemia and reperfusion (IR) injury on excitation-contraction coupling in hearts. We 1) quantified the phase-space relationship between simultaneously measured myoplasmic Ca2+ concentration ([Ca2+]) and isovolumetric left ventricular pressure (LVP) using indexes of loop area, orientation, and position; and 2) quantified cooperativity by linearly modeling the phase-space relationship between [Ca2+] and rate of LVP development in intact hearts during administration of positive inotropic drugs before and after global IR injury. Unpaced, isolated guinea pig hearts were perfused at a constant pressure with Krebs-Ringer solution (37 degrees C, 1.25 mM CaCl2). [Ca2+] was measured ratiometrically by indo 1 fluorescence by using a fiber-optic probe placed at the left ventricular free wall. LVP was measured by using a saline-filled latex balloon and transducer. Drugs were infused for 2 min, 30 min before, and for 2 min, 30 min after 30-min global ischemia. IR injury worsened Ca2+-contraction coupling, as seen from decreased orientation and repositioning of the loop rightward and downward and reduced cooperativity of contraction and relaxation with or without drugs. Dobutamine (4 microM) worsened, whereas dopamine (8 microM) improved Ca2+-contraction coupling before and after IR injury. Dobutamine and dopamine improved cooperativity of contraction and relaxation after IR injury, whereas only dopamine increased cooperativity of relaxation before IR injury. Digoxin (1 microM) improved Ca2+-contraction coupling and cooperativity of contraction after but not before ischemia. Levosimendan (1 microM) did not alter Ca2+-contraction coupling or cooperativity, despite producing concomitant increases in contractility, relaxation, and Ca2+ flux before and after ischemia. Dynamic indexes based on LVP-[Ca2+] diagrams (area, shape, position) can be used to identify and measure alterations in Ca2+-contraction coupling during administration of positive inotropic drugs in isolated hearts before and after IR injury.