Stimuli that possess inherently rewarding or aversive qualities elicit emotional responses and also induce learning by imparting valence upon neutral sensory cues. Evidence has accumulated ...implicating the amygdala as a critical structure in mediating these processes. We have developed a genetic strategy to identify the representations of rewarding and aversive unconditioned stimuli (USs) in the basolateral amygdala (BLA) and have examined their role in innate and learned responses. Activation of an ensemble of US-responsive cells in the BLA elicits innate physiological and behavioral responses of different valence. Activation of this US ensemble can also reinforce appetitive and aversive learning when paired with differing neutral stimuli. Moreover, we establish that the activation of US-responsive cells in the BLA is necessary for the expression of a conditioned response. Neural representations of conditioned and unconditioned stimuli therefore ultimately connect to US-responsive cells in the BLA to elicit both innate and learned responses.
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•ChR2 was expressed in unconditioned stimulus (US) representations in the BLA•Activation of US representations elicits innate responses and drives learning•Inhibition of US representations prevents the expression of learned behavior•Conditioned stimuli activate US representations in BLA to elicit learned behavior
Neurons in the basolateral amygdala that mediate responses to intrinsically rewarding or aversive stimuli also elicit learned responses, indicating that associative learning is funneled through innate behavioral circuits to assign positive or negative emotions to neutral sensory stimuli.
Controlling protein function through posttranslational manipulations has emerged as an attractive complementary technology to existing genetic systems. Often these methods involve developing ...pharmacological agents to probe protein function without the need to generate a unique compound for each protein family. One common strategy uses small molecules that act as chemical inducers of dimerization by mediating the interaction of two proteins. Herein we report the use of a chemical inducer of dimerization for the development of a posttranslational technology for the manipulation of protein function. This system, split ubiquitin for the rescue of function (SURF), places the complementation of genetically split ubiquitin under the control of rapamycin-induced dimerization of FK506-binding protein and FKBP12-rapamycin-binding protein. Before complementation a "degron" dooms a protein of interest for destruction by the proteasome. Addition of rapamycin results in a proteolytic shunt away from degradation by inducing ubiquitin complementation and cleavage of the protein of interest from the degron. Importantly, the native protein is rescued. We characterized this system with firefly luciferase and went on to apply it to members of three important classes of proteins: proteases (caspase-3), kinases (v-Src), and transcription factors (Smad3). This general strategy should allow for inducible rescue of a variety of proteins in such a way that their native structure and function are maintained.
Control over the timing, location and level of protein activity in vivo is crucial to understanding biological function. Living systems are able to respond to external and internal stimuli rapidly ...and in a graded fashion by maintaining a pool of proteins whose activities are altered through post-translational modifications. Here we show that the process of protein trans-splicing can be used to modulate enzymatic activity both in cultured cells and in Drosophila melanogaster. We used an optimized conditional protein splicing system to rapidly trigger the in vivo ligation of two inactive fragments of firefly luciferase in a tunable manner. This technique provides a means of controlling enzymatic function with greater speed and precision than with standard genetic techniques and is a useful tool for probing biological processes.
Understanding brain function requires knowing both how neural activity encodes information and how this activity generates appropriate responses. Electrophysiological, imaging and immediate early ...gene immunostaining studies have been instrumental in identifying and characterizing neurons that respond to different sensory stimuli, events and motor actions. Here we highlight approaches that have manipulated the activity of physiologically classified neurons to determine their role in the generation of behavioural responses. Previous experiments have often exploited the functional architecture observed in many cortical areas, where clusters of neurons share response properties. However, many brain structures do not exhibit such functional architecture. Instead, neurons with different response properties are anatomically intermingled. Emerging genetic approaches have enabled the identification and manipulation of neurons that respond to specific stimuli despite the lack of discernable anatomical organization. These approaches have advanced understanding of the circuits mediating sensory perception, learning and memory, and the generation of behavioural responses by providing causal evidence linking neural response properties to appropriate behavioural output. However, significant challenges remain for understanding cognitive processes that are probably mediated by neurons with more complex physiological response properties. Currently available strategies may prove inadequate for determining how activity in these neurons is causally related to cognitive behaviour.
Understanding brain function requires knowing both how neural activity encodes information and how this activity generates appropriate responses. Electrophysiological, imaging and immediate early ...gene immunostaining studies have been instrumental in identifying and characterizing neurons that respond to different sensory stimuli, events and motor actions. Here we highlight approaches that have manipulated the activity of physiologically classified neurons to determine their role in the generation of behavioural responses. Previous experiments have often exploited the functional architecture observed in many cortical areas, where clusters of neurons share response properties. However, many brain structures do not exhibit such functional architecture. Instead, neurons with different response properties are anatomically intermingled. Emerging genetic approaches have enabled the identification and manipulation of neurons that respond to specific stimuli despite the lack of discernable anatomical organization. These approaches have advanced understanding of the circuits mediating sensory perception, learning and memory, and the generation of behavioural responses by providing causal evidence linking neural response properties to appropriate behavioural output. However, significant challenges remain for understanding cognitive processes that are probably mediated by neurons with more complex physiological response properties. Currently available strategies may prove inadequate for determining how activity in these neurons is causally related to cognitive behaviour.
The σ factors are the key regulators of bacterial transcription initiation. Through direct read-out of promoter DNA sequence, they recruit the core RNA polymerase to sites of initiation, thereby ...dictating the RNA polymerase promoter-specificity. The group 1 σ factors, which direct the vast majority of transcription initiation during log phase growth and are essential for viability, are autoregulated by an N-terminal sequence known as σ1.1. We report the solution structure of Thermotoga maritima σA σ1.1. We additionally demonstrate by using chemical crosslinking strategies that σ1.1 is in close proximity to the promoter recognition domains of σA. We therefore propose that σ1.1 autoinhibits promoter DNA binding of free σA by stabilizing a compact organization of the σ factor domains that is unable to bind DNA.
SUMMARYIn severe pump failure with hypotension complicating acute myocardial infarction, dopamine has been useful in raising arterial pressure by increasing myocardial performance and augmenting ...peripheral resistance. Once adequate blood pressures are obtained, vasodilators may be used to reduce peripheral resistance and thus enhance pump performance. Accordingly, the hemodynamic effects of dopamine and nitroprusside administration were monitored in eight patients who developed hypotension following an acute myocardial infarction. With dopamine therapy alone, mean arterial pressure averaged 84 $pM 3.6 mm Hg and mean left ventricular filling pressure 32 $pM 7.9 mm Hg. The addition of nitroprusside, at doses ranging from 0.5 to 1.6 $mUg/kg/min, resulted in a decrease in arterial pressure to 75 $pM 2.4 mm Hg (p < 0.01) and in left ventricular filling pressure to 23 $pM 7.2 mm Hg (p < 0.001). Cardiac index increased modestly from 1.81 $pM 0.61 to 2.01 $pM 0.60 liters/min/m (p < 0.05), while systemic vascular resistance fell from 1,967 $pM 707 to 1,586 $pM 634 dynes-sec-cm (p < 0.01). Heart rate did not change significantly. Seven of eight patients died in the hospital within 4 days of admission. It appears that despite a beneficial hemodynamic response effected by combined dopamine-nitroprusside administration, the prognosis of this group of patients may not be favorably altered because of the extensive destruction of myocardium.
We have been attempting to develop a consistently reliable internal control to assure the effectiveness of the 5-fluorodeoxyuridine (FUdR) fragile-X fra(X) induction system. We carried out a ...systematic study of whole-blood specimens cultured from 56 individuals from two different laboratories. An analysis of nearly 9,000 cells demonstrated: (1) the importance of establishing baseline levels of fragile sites in each laboratory, and (2) that a combination of common fragile sites (different for each laboratory) could serve as a consistently reliable indicator of the effectiveness of the FUdR fra(X) induction system. It was suggested that a non-FUdR culture(s) should be incorporated into a laboratory's fra(X)-screening protocol, so that if there are any doubts about the effectiveness of the FUdR system a comparison to background or spontaneously occurring fragile sites can be made within the laboratory. Repeat cultures are recommended where no increase in common fragile-site frequency is observed in the FUdR induction system, and where fra(X) was strongly suspected but not found. In addition, the necessity of using more than one fra(X) induction system in whole-blood cultures was demonstrated, including the effectiveness of an FUdR/excess thymidine double-induction system. Finally, 2 cases of apparent mosaicism for Klinefelter syndrome in fra(X) individuals were observed.
Three cases of distal duplication 14q are presented. The first two cases are cousins in a kindred segregating a balanced translocation t(14;18)(q31;q23). The third case resulted from a maternal ...translocation t(14;18)(q24;p11). By review of these cases and those previously reported, a distal duplication 14q syndrome is further delineated. Common features include postnatal growth retardation, mental retardation, hypotonia, microcephaly, slanted palpebral fissures, ocular hypertelorism, sparse eyelashes and eyebrows, nasal dysmorphism, tented lip, micrognathia, posteriorly rotated ears, and minor skeletal anomalies.
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