ABSTRACT
Underactive bladder or detrusor underactivity (DU) is defined as a reduction of contraction strength or duration of the bladder wall. Despite the serious healthcare implications of DU, there are limited solutions for affected individuals. A flexible 3D printed implantable device driven by shape memory alloys (SMA) actuators is presented here for the first time to physically contract the bladder to restore voluntary control of the bladder for individuals suffering from DU. This approach is used initially in benchtop experiments with a rubber balloon acting as a model for the rat bladder to verify its potential for voiding, and that the operating temperatures are safe for the eventual implantation of the device in a rat. The device is then implanted and tested on an anesthetized rat, and a voiding volume of more than 8% is successfully achieved for the SMA-based device without any surgical intervention or drug injection to relax the external sphincter.
ABSTRACT
Long-term potentiation (LTP) of excitatory synapses on ventral tegmental area (VTA) dopamine (DA) cells is thought to play an important role in mediating some of the behavioral effects of drugs of abuse yet little is known about its underlying mechanisms. We find that spike timing-dependent LTP (STD LTP) in VTA DA cells is absent in slices prepared from mice previously administered cocaine, suggesting that cocaine-induced LTP and STD LTP share underlying mechanisms. This form of STD LTP is dependent on NMDA receptor (NMDAR) activation and a rise in postsynaptic calcium but surprisingly was not affected by an inhibitor of calcium/calmodulin-dependent protein kinase II (CaMKII). It was blocked by antagonists of conventional isoforms of PKC, whereas activation of protein kinase C (PKC) using a phorbol ester enhanced synaptic strength. These results suggest that NMDAR-mediated activation of PKC, but not CaMKII, is a critical trigger for LTP in VTA DA cells.
Subject(s)
Dopamine/metabolism , Long-Term Potentiation/physiology , Neurons/physiology , Protein Kinase C/physiology , Ventral Tegmental Area/cytology , Animals , Cocaine/pharmacology , Dopamine Uptake Inhibitors/pharmacology , Electric Stimulation/methods , Enzyme Inhibitors/pharmacology , Excitatory Amino Acid Antagonists/pharmacology , Excitatory Postsynaptic Potentials/drug effects , Excitatory Postsynaptic Potentials/physiology , Excitatory Postsynaptic Potentials/radiation effects , Female , In Vitro Techniques , Long-Term Potentiation/drug effects , Long-Term Potentiation/radiation effects , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neurons/radiation effects , Patch-Clamp Techniques/methods , Phorbol Esters/pharmacology , Time Factors , Valine/analogs & derivatives , Valine/pharmacologySubject(s)
Neurons, Afferent/physiology , Receptors, G-Protein-Coupled/metabolism , Receptors, Odorant/metabolism , Smell/physiology , Animals , Genetic Vectors , Humans , Ligands , Models, Biological , Neurons, Afferent/metabolism , Protein Binding , Signal Transduction , Vomeronasal Organ/metabolismABSTRACT
The identification of the chemical structure of an odorant by the vertebrate olfactory system is thought to occur through the combinatorial activity from multiple receptors, each tuned to recognize different chemical features. What are the molecular determinants underlying the selectivity of individual odorant receptors for their cognate ligands? To address this question, we performed molecular modeling and site-directed mutagenesis on the ligand-binding region of two orthologous amino acid odorant receptors belonging to the "C family" of G-protein-coupled receptors in goldfish and zebrafish. We identified the critical ligand-receptor interactions that afford ligand binding as well as selectivity for different amino acids. Moreover, predictions regarding binding pocket structure allowed us to alter, in a predictable manner, the receptor preferences for different ligands. These results reveal how this class of odorant receptor has evolved to accommodate ligands of varying chemical structure and further illuminate the molecular principles underlying ligand recognition and selectivity in this family of chemosensory receptors.
Subject(s)
Amino Acids/metabolism , Receptors, Odorant/chemistry , Zebrafish Proteins/chemistry , Amino Acids/chemistry , Animals , Arginine/chemistry , Arginine/metabolism , Binding Sites , Calcium/analysis , Cell Line/chemistry , DNA, Complementary/genetics , Gene Library , Glycine/chemistry , Goldfish/genetics , Humans , Kidney/chemistry , Kidney/cytology , Ligands , Models, Molecular , Monte Carlo Method , Mutagenesis, Site-Directed , Protein Binding , Protein Conformation , Protein Structure, Tertiary , Receptors, Odorant/genetics , Receptors, Odorant/metabolism , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/metabolism , Species Specificity , Structure-Activity Relationship , Substrate Specificity , Zebrafish Proteins/geneticsABSTRACT
There are many sources of systematic variation in cDNA microarray experiments which affect the measured gene expression levels (e.g. differences in labeling efficiency between the two fluorescent dyes). The term normalization refers to the process of removing such variation. A constant adjustment is often used to force the distribution of the intensity log ratios to have a median of zero for each slide. However, such global normalization approaches are not adequate in situations where dye biases can depend on spot overall intensity and/or spatial location within the array. This article proposes normalization methods that are based on robust local regression and account for intensity and spatial dependence in dye biases for different types of cDNA microarray experiments. The selection of appropriate controls for normalization is discussed and a novel set of controls (microarray sample pool, MSP) is introduced to aid in intensity-dependent normalization. Lastly, to allow for comparisons of expression levels across slides, a robust method based on maximum likelihood estimation is proposed to adjust for scale differences among slides.
