Biomedical subjects
Publications and source records attributed to D O Carpenter.
We describe an apparatus for immobilization of the rattlesnake head for brain experimentation. Its design also allows its use as a stereotaxic instrument for electrophysiological studies employing microelectrodes. Attachment of the head is at the two orbital ridges and the hard palate. The bregma provides a zero reference for the frontal, lateral, and depth coordinates.
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MCF-7 cell growth is normally dependent upon estrogen, but if cells are maintained in serum-free medium estrogen inhibits cell growth with an IC50 of about 1 nM. Cells adapted to serum-free medium have approximately the same levels of estrogen receptor mRNA as control cells that are stimulated by estrogen. We have identified two serum components, one estrogen dependent and one not, that appear to be responsible for the inhibition of growth by estrogen. Our observations are consistent with the presence of a plasma membrane receptor which negatively regulates MCF-7 cell growth, and that can be inhibited by a serum protein that binds estrogen.
We have described two different proteins that mediate MCF-7 cell growth inhibition (1). One is estrogen-dependent and is a heterodimer consisting of a heavy (54 kDa) and light (29 kDa) chain. This protein is distinct from cortisol-binding globulin (CBG), sex hormone-binding protein (SBP or SHBG) and albumin (HSA). It may be fetal steroid binding protein (FSBP). The other is not estrogen dependent, but may be related to the estrogen-dependent protein. Since these are large proteins they must be acting at the level of the plasma membrane, not traditional intracellular estrogen receptors, and inhibit MCF-7 cell growth.
It was shown that acetylcholine and gamma-aminobutyric acid at extremely low concentrations, which do not activate receptor-binding ionic channels, modulate the activity of NA-K-pump and Na:Ca exchange, the intracellular level of cAMP and chemosensitivity of the neuronal membrane. These data provide a new evidence in support for Koshtoiants's enzymochemical hypothesis in which synaptic transmitters are considered to be metabolic modulators for postsynaptic neurones.
We have compared the effects of Pb2+ on voltage-dependent calcium channels of the marine mollusc Aplysia, studied with a two electrode voltage clamp, with those on calcium channels in cultured rat dorsal root ganglion (DRG) neurons studied with whole cell patch clamp. In both preparations Pb2+ was a potent blocker of calcium channel currents at concentrations that did not significantly affect potassium and sodium currents. The blockade was concentration dependent and the percentage of blockade was reduced when the concentration of the charge carrier was elevated. In Aplysia the threshold Pb2+ concentration was about 1 microM, and the Hill coefficient near 1.0 under all conditions. Pb2+ did not significantly change inactivation but shifted the voltage dependence of activation to hyperpolarized voltages in a dose-dependent manner. The blockade of calcium currents by Pb2+ was highly voltage dependent and increased with depolarization. Rat dorsal root ganglion cells exhibit three different types of voltage-dependent calcium channels (N, L and T) which can be distinguished by the potential at which the channel activates or inactivates and by their sensitivity to pharmacologic antagonists. The IC50 for blockade of the L current was 1.03 microM with a Hill slope of 1.15. Currents elicited by voltage steps which activate N plus L currents had an IC50 of 0.64 microM and a Hill slope of 1.16. T currents were less sensitive, having an IC50 of 6 microM. Sodium and potassium currents were relatively unaffected in both preparations at concentrations at which the calcium channel was blocked more than 60% (1 microM or 200 microM respectively). The blockade in DRG neurons was less voltage-dependent and reversible than that in the invertebrate model system. These observations indicate that Pb2+ is a potent, reversible and selective blocker of voltage-dependent calcium channels at low concentrations.
Total dissolved Pb was measured in a number of commonly used physiological solutions by electrothermal atomization atomic absorption spectrometry (ETA-AAS). In HEPES-buffered solutions (pH 7.30) the concentration of total Pb in solution ("measured" Pb) was only 77% of nominal Pb up to 20 microM added Pb, where experiments were undertaken at room temperature (22 +/- 1 degree C). However, test solutions equilibrated at 37 +/- 1 degree C contained 99% of added Pb up to 2 microM. Above this nominal concentration, percentage recoveries dropped to approximately 72% at a nominal concentration of 20 microM. Tris-buffered artificial seawater (CaASW) solutions (pH 7.60) contained more dissolved Pb compared to HEPES-buffered solutions at 22 degrees C. However, increasing the calcium concentration in ASW appeared to increase precipitation of PbCl2. Concentration-corrected dose-response relationships were plotted from previously published data on the effects of Pb on voltage-activated calcium channels of rat dorsal root ganglion (DRG) cells and Aplysia neurons. The plots suggest that the inhibitory effects of Pb on rat DRG cells may prevail at concentrations of Pb even lower than reported previously when measurements were made at 22 degrees C. However, increasing the temperature to 37 degrees C resulted in closer agreement between measured and nominal dose-response curves. The measured dose-response curves for the Tris-buffered ASW solutions closely followed those of the nominal up to 200 microM Pb. In ASW solutions containing 40 mM calcium, PbCl2 was precipitated at Pb concentrations greater than 200 microM.