Bovine ovarian non-genomic progesterone binding sites: presence in follicular and luteal cell membranes

We have shown recently that the bovine corpus luteum (CL) possesses specific luteal cell surface membrane binding sites for progesterone. We have now confirmed and extended these observations to compare the subcellular distribution of these binding sites in developing, mature and regressing CL. The median buoyant densities of luteal progesterone binding sites from early-, mid- and late-luteal phase CL were similar (though three of five density profiles for late-luteal phase CL showed association of steroid binding with a fraction with a lower density), and clearly resolved from nuclear, mitochondrial, lysosomal, peroxisomal, Golgi-endoplasmic reticulum-lysosomal and smooth endoplasmic reticulum markers. Specific binding of [3H]progesterone overlapped with the distributions of 5'-nucleotidase and luteinizing hormone receptor (luteal cell surface membrane markers) in both control and digitonin-treated gradients at all stages of the luteal phase. Since steroidogenic 'large luteal' and 'small luteal' cells of the CL are derived from the granulosa cells (GC) and theca of the preovulatory follicle, we also investigated whether similar receptors were present in the follicle, and describe for the first time specific membrane binding sites for progesterone in purified GC and thecal membranes from healthy bovine follicles of different sizes. Specific binding increased linearly with GC and thecal membrane protein concentration; however, it was detectable only when digitonin was included in the binding incubation. Binding sites were specific for progesterone; unlabelled progesterone competed for [3H]progesterone binding at low concentrations (IC50, 35 and 31 nmol/l) compared with testosterone (IC50, 905 and 870 nmol/l) and delta4-androstenedione (IC50, 1050 and 660 nmol/l) for GC and thecal receptors respectively. In contrast, oestradiol, oestrone, pregnenolone, cortisol, cholesterol, and a genomic progesterone receptor antagonist, RU486, competed poorly. Steroid binding was present in GC and thecal membranes of follicles of all sizes, but [3H]progesterone binding to GC membranes decreased significantly with increasing follicle size (P<0.02), perhaps indicating developmental regulation of GC membrane non-genomic progesterone receptors in the preovulatory bovine follicle. We suggest that these membrane steroid receptors may be involved in the autocrine/paracrine regulation of follicular function by progesterone.     

Cellular mechanisms for developmental toxicity of chlorpyrifos: targeting the adenylyl cyclase signaling cascade

Developmental neurotoxicity caused by chlorpyrifos exposure is generally thought to target cholinesterase but chlorpyrifos may also act on cellular intermediates, such as adenylyl cyclase, that serve global functions in the coordination of cell development. In the current study, neonatal rats were exposed to apparently subtoxic doses of chlorpyrifos (no weight loss, no mortality) either on Postnatal Days 1-4 or on Postnatal Days 11-14, and the effects on components of the adenylyl cyclase cascade were evaluated in brain regions that are enriched (forebrain) or sparse (cerebellum) in cholinergic innervation, as well as in a nonneural tissue (heart). In all three, chlorpyrifos evoked deficits in multiple components of the adenylyl cyclase cascade: expression and activity of adenylyl cyclase itself, functioning of G-proteins that link neurotransmitter and hormone receptors to cyclase activity, and expression of neurotransmitter receptors that act through this cascade. Disruption of signaling function was not restricted to transduction of cholinergic signals but rather extended to adrenergic signals as well. In most cases, the adverse effects were not evident during the immediate period of chlorpyrifos administration, but appeared after a delay of several days. These results suggest that chlorpyrifos can affect cell development by altering the activity and reactivity of the adenylyl cyclase signaling cascade, a major control point for trophic regulation of cell differentiation. The effects are not restricted to cholinergic targets, nor even to the central nervous system. Hence, disruption of cell development by chlorpyrifos is likely to be more widespread than previously thought.     

Rapid identification of protein phosphatase 1-binding proteins by mixed peptide sequencing and data base searching. Characterization of a novel holoenzymic form of protein phosphatase 1

Microcystin-affinity chromatography was used to purify 15 protein phosphatase 1 (PP1)-binding proteins from the myofibrillar fraction of rabbit skeletal muscle. To reduce the time and amount of material required to identify these proteins, proteome analysis by mixed peptide sequencing was developed. Proteins are resolved by SDS-polyacrylamide gel electrophoresis, electroblotted to polyvinylidene fluoride membrane, and stained. Bands are sliced from the membrane, cleaved briefly with CnBr, and applied without further purification to an automated Edman sequencer. The mixed peptide sequences generated are sorted and matched against the GenBank using two new programs, FASTF and TFASTF. This technology offers a simple alternative to mass spectrometry for the subpicomolar identification of proteins in polyacrylamide gels. Using this technology, all 15 proteins recovered in PP-1C affinity chromatography were sequenced. One of the proteins, PP-1bp55, was homologous to human myosin phosphatase, MYPT2. A second, PP-1bp80, identified in the EST data bases, contained a putative PP-1C binding site and a nucleotide binding motif. Further affinity purification over ATP-Sepharose isolated PP-1bp80 in a quaternary complex with PP-1C and two other proteins, PP-1bp29 and human p20. Recombinant PP-1bp80 also bound PP-1C and suppressed its activity toward a variety of substrates, suggesting that the protein is a novel regulatory subunit of PP-1.     

A two-stage algorithm for identification of nonlinear dynamic systems

This paper investigates the two-stage stepwise identification for a class of nonlinear dynamic systems that can be described by linear-in-the-parameters models, and the model has to be built from a very large pool of basis functions or model terms. The main objective is to improve the compactness of the model that is obtained by the forward stepwise methods, while retaining the computational efficiency. The proposed algorithm first generates an initial model using a forward stepwise procedure. The significance of each selected term is then reviewed at the second stage and all insignificant ones are replaced, resulting in an optimised compact model with significantly improved performance. The main contribution of this paper is that these two stages are performed within a well-defined regression context, leading to significantly reduced computational complexity. The efficiency of the algorithm is confirmed by the computational complexity analysis, and its effectiveness is demonstrated by the simulation results.     

Apollonius tenth problem via radius adjustment and Möbius transformations

The Apollonius Tenth Problem, as defined by Apollonius of Perga circa 200 B.C., has been useful for various applications in addition to its theoretical interest. Even though particular cases have been handled previously, a general framework for the problem has never been reported. Presented in this paper is a theory to handle the Apollonius Tenth Problem by characterizing the spatial relationship among given circles and the desired Apollonius circles. Hence, the given three circles in this paper do not make any assumption regarding on the sizes of circles and the intersection/inclusion relationship among them. The observations made provide an easy-to-code algorithm to compute any desired Apollonius circle which is computationally efficient and robust.     

Complete mobile email management

Years have lapsed since the inauguration of the first Wireless Application Protocol (WAP) application. One of the main reasons for the lukewarm response so far may be attributed to the lack of a “Killer” application. Many companies and developers have attempted to replicate email services (an Internet Killer appplication) within the WAP realm, but till now, no significant success has been reported. This paper will first discuss on the key functions which are lacking in the present WAP-email service, and which are arguably necessary if a much larger user base is to be attracted to these services. A new system will be proposed which can enable the user to send files residing in a remote personal computer (PC) as attachments to his emails using WAP-enabled mobile devices. The implementation of the system will be presented in details. Issue relating to the connection to various types of email servers and the security of the system will also be addressed in the paper.     

Hardware and software architecture for power quality analysis

This paper presents a novel concept for power quality hardware and software architecture. Software tools related to detection, classification and characterization of power quality events can be integrated into a Distribution Control Center in which the interconnection between different subsystems is being carried out through the Internet allowing flexibility to the system. This system is implemented using wavelet analysis applied in all proposed algorithms. Different power quality events are taken as examples to illustrate the capabilities of the proposed method. The obtained results reveal that the proposed architecture is feasible to be implemented as an integral part of a control center system.The proposed system has the property of fast and accurate detection and classification of any power quality disturbance event and introduces a new PQ index determination that allows characterizing any type of disturbance including the non-periodic signals.