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The venoms of predatory cone snails harbor a rich repertoire of peptide toxins that are valuable research tools, but recently have also proven to be useful drugs. Among the conotoxins with several disulfide bridges, the O-superfamily toxins are characterized by a conserved cysteine knot pattern: C-C-CC-C-C. While ω-conotoxins and κ-conotoxins block Ca2+ and K+ channels, respectively, the closely related δ- and μO-conotoxins affect voltage-gated Na+ channels (Nav channels). δ-conotoxins mainly remove the fast inactivation of Nav channels and, thus, functionally resemble long-chain scorpion α-toxins. μO-conotoxins are functionally similar to μ-conotoxins, since they inhibit the ion flow through Nav channels. Recent results from functional and structural assays have gained insight into the underlying molecular mechanisms. Both types of toxins are voltage-sensor toxins interfering with the voltage-sensor elements of Nav channels. [ABSTRACT FROM AUTHOR]

Large-conductance Ca2+-dependent K+ (BKCa) channels are activated by intracellular Ca2+ and membrane depolarization in an allosteric manner. We investigated the pharmacological and biophysical characteristics of a BKCa-type K+ channel in androgen-dependent LNCaP (lymph node carcinoma of the prostate) cells with novel functional properties, here termed BKL. K+ selectivity, high conductance, activation by Mg2+ or NS1619, and inhibition by paxilline and penitrem A largely resembled the properties of recombinant BKCa channels. However, unlike conventional BKCa channels, BKL channels activated in the absence of free cytosolic Ca2+ at physiological membrane potentials; the half-maximal activation voltage was shifted by about −100 mV compared with BKCa channels. Half-maximal Ca2+-dependent activation was observed at 0.4 μ M for BKL (at −20 mV) and at 4.1 μ M for BKCa channels (at +50 mV). Heterologous expression of hSlo1 in LNCaP cells increased the BKL conductance. Expression of hSlo-β 1 in LNCaP cells shifted voltage-dependent activation to values between that of BKL and BKCa channels and reduced the slope of the P open (open probability)-voltage curve. We propose that LNCaP cells harbor a so far unknown type of BKCa subunit, which is responsible for the BKL phenotype in a dominant manner. BKL-like channels are also expressed in the human breast cancer cell line T47D. In addition, functional expression of BKL in LNCaP cells is regulated by serum-derived factors, however not by androgens. [ABSTRACT FROM AUTHOR]

Abstract Shaker B potassium channels undergo rapid N-type and slow C-type inactivation. While N-type inactivation is supposed to be mediated by occlusion of the pore by the N-terminal protein structure, the molecular mechanisms leading to C-type inactivation are less well understood. Considering N-type inactivation as a model for a protein conformational transition, we investigated inactivation of heterologously expressed Shaker B potassium channels and mutants thereof, showing various degrees of C-type inactivation, under high hydrostatic (oil) pressure. In addition to the derived apparent activation and reaction volumes (change V), experiments at various temperatures yielded estimates for enthalpic (change H) and entropic (T change S) contributions. N-type inactivation was accelerated by increasing temperature and slowed by high hydrostatic pressure yielding at equilibrium change H=76 kJ/mole, T change S = 82 kJ/mole, and change V = 0.18 nm[sup 3] indicating that the transition to the N-type inactivated state is accompanied by an increase in volume and a decrease in order. N-terminally deleted SH change 6-46 constructs with additional mutations at either position 449 or 463 were used to investigate C-type inactivation. In particular at high temperatures, inactivation occurred in two phases indicating more than one process. At equilibrium the following values were estimated for the major inactivation component of mutant Sh change 6-46 T449A: change H=-64 kJ/mole, T change S=-60 kJ/mole, and change V=-0.25 nm[sup 3], indicating that the C-type inactivated state occupies a smaller volume and is more ordered than the noninactivated state. Thus, hydrostatic pressure affects N- and C-type inactivation in opposite ways. [ABSTRACT FROM AUTHOR]

One of the major sources of despair for the investigator who hopes to explore properties of the Thematic Apperception Test (TAT) is the deadly business of collecting this kind of data. Clinician and research worker alike find this feature of the test a barrier to learning more about the instrument and thus eventually about human behavior. To some extent the time involved in administering the test may be seen as a matter of inconvenience for the sake of a valued end since the potential power of the instrument is certainly related to the richness of the material it elicits, and this in turn is linked to its awkwardness as a data-collection procedure. However, if the process of data collection can be streamlined without seriously impairing the test's sensitivity, this will clearly represent a valuable contribution. The study reported here addresses itself to the problem of finding out to what extent findings secured from the TAT under conditions of group administration may be considered equivalent to those secured with individual administration. Related to this is the question of whether there is any striking superiority of one method of administration over the other.

Phosphoinositid 3-Kinase γ (PI3Kγ), the only known member of class Ib PI3K's, is activated by G-Protein coupled receptors (GPCRs), producing the second messenger phosphatidylinositol-3,4,5-trisphosphate (PIP3). In addition, PI3Kγ possesses an intrinsic protein kinase activity and might express a scaffold function in the signaling network of different cells. PI3Kγ is expressed in hematopoietic cells acting as a main mediator of proliferation, differentiation, migration and survival of leukocytes. Recent studies expand these well-established cell specific functions of PI3Kγ to other cell types like cardiomyocytes. In heart PI3Kγ exhibits a negative regulatory function on the contractility by controlling the intracellular camp-level in concert with Phosphodiesterase 3B (PDE3B). In the present study we describe for the first time PI3Kγ expression and functional pattern in cell types of neuronal origin. PI3Kγ-protein is expressed in small diameter neurons of dorsal root ganglia (DRG), which form widely unmyelinated C-fibers into the body's periphery. These primary sensory neurons exhibit nociceptive functions and can lead to a slow, dull and long lasting pain perception. These neurons are heavily involved in the processes of neuropathic and inflammatory pain accompanied by molecular modifications in the interplay of extra- and intracellular milieus -- the playground of PI3Kγ. Now we take advantage of the PI3Kγ-knockout mice to evaluate the impact of PI3Kγ in behavioral studies and primary dorsal root ganglia cultures. Analysis of nociceptive behaviors and studies on DRG-cultures suggest an involvement of the signaling protein in the μ-opioid-receptor path. An important regulation of the opioid-receptor system is the sensitization and desensitization of the receptors. Effects of different μ-receptor-agonists point to a differential involvement of PI3Kγ in antinociceptive signaling in primary sensory neurons. [ABSTRACT FROM AUTHOR]

Abstract Voltage-activated, Ca2+ -sensitive K+ channels (BK or maxi K,Ca channels) play a major role in the control of neuronal excitability. We have cloned pSlo, the BK channel α subunit of the cockroach Periplaneta americana . The amino acid sequence of pSlo shows 88% identity to dSlo from Drosophila . There are five alternatively spliced positions in pSlo showing differential expression in various tissues. A pSlo-specific antibody prominently stained the octopaminergic dorsal unpaired median (DUM) neurons and peptidergic midline neurons in Periplaneta abdominal ganglia. HEK293 cells expressing pSlo exhibit K+ channels of 170 pS conductance. They have a tendency for brief closures, exhibit subconductance states and show slight inward rectification. Activation kinetics and voltage dependence are controlled by cytoplasmic [Ca2+ ]. In contrast to dSlo, pSlo channels are sensitive to charybdotoxin and iberiotoxin. Mutagenesis at two positions (E254 and Q285) changed blocking efficacy of charybdotoxin. In contrast to pSlo expressed in HEK293 cells, native IbTx-sensitive K,Ca currents in DUM and in peptidergic neurons, exhibited rapid, partial inactivation. The fast component of the K,Ca current partly accounts for the repolarization and the early after-hyperpolarization of the action potential. By means of Ca2+ -induced repolarization, BK channels may reduce the risk of Ca2+ overload in cockroach neurons. Interestingly, the neurons expressing pSlo were also found to express taurine, a messenger that is likely to limit overexcitation by an autocrine mechanism in mammalian central neurons. [ABSTRACT FROM AUTHOR]