Effects of progestins on the proliferation of estrogen-dependent human breast cancer cells under growth factor-defined conditions.

The effect was studied of four different synthetic progestins (Org 30659, gestodene, 3-ketodesogestrel and levonorgestrel) on the proliferation of the 17 beta estradiol (E2)-dependent human breast cancer cell line MCF7. All progestins were found to stimulate proliferation, but only at high pharmacological dosages. Moreover, like estrogens the progestins at high concentrations synergistically stimulated MCF7 cell proliferation together with low concentrations of insulin. This stimulatory effect could be blocked by antiestrogens, but not by antiglucocorticoids and antiprogestins. This suggests that growth stimulation by these progestins (or their metabolites) occurs through crossreaction with the E2 receptor (ER). This is confirmed by the observation that the strong synthetic progestin Org 2058 does not stimulate proliferation. The absence of a progesterone receptor (PR)-mediated growth response seems not to be due to aberrant PR expression in these cells; 27,000 receptors (Kd 1.7 x 10(-10)M) per cell were present under growth-assay conditions. Growth stimulation by E2 in the absence or presence of insulin, is slightly inhibited or unaffected by the progestins, respectively. Our data do not support a role for the recently identified gestodene binding sites [Colletta et al., J. Steroid Biochem. 33 (1989) 1055-1061] in mediating gestodene effects on breast cancer cells: gestodene and 3-ketodesogestrel, a compound that does not bind to these gestodene binding sites, showed a similar biological activity. The effects of the progestins on the MCF7 breast cancer cell line, indicate that the use of these compounds at very high concentrations may be unfavourable, but do not support a role for them in directly stimulating breast tumor proliferation at the low progestin concentration which are reached in the serum in oral contraceptive users.     

Effects of Somatosensory Stimulation on Dream Content in Gymnasts and Control Participants: Evidence of Vestibulomotor Adaptation in REM Sleep

Somatosensory stimulation of the leg muscles in REM sleep appears to perturb virtual orientation in dream experiences. According to our model of vestibulomotor adaptation (Sauvageau, Nielsen, & Montplaisir, 1996), the dreaming mind attempts to compensate for such destabilizing stimulation by increasing eye movement activity or by modifying dream content, among other possible reactions. Effective compensation may be more easily achieved by participants who are adapted to the disruptive stimulation or who possess highly developed vestibulomotor skills. To examine this possibility, we studied the effects of Somatosensory stimulation on the dreams of 6 gymnasts and 6 control participants aged 9 to 16 years. Results provide some support for the expectations that 1) imposed Somatosensory information is processed by the central nervous system in REM sleep, 2) unilateral stimulation induces an upset in virtual orientation, 3) gymnasts are more resistant to these disruptive effects of stimulation than are control participants, and 4) because of long-term adaptation, the dream content of gymnasts does not differ markedly from that of controls. Though preliminary and in need of replication, the findings are compatible with the notion that the developed vestibular skills of gymnasts protects them to some extent from the effects of a disruptive Somatosensory stimulus during sleep.     

Effect of dry immersion in combination with stimulation of foot support zones upon muscle force-velocity characteristics

The complex of motor disturbances arising under conditions of real and simulated microgravity that include decrease of contractile characteristics of postural muscles is likely to be a result of withdrawal of support stimuli. Artificial stimulation of support zones of feet is shown to diminish partially or prevent completely the negative effects of microgravity on the motor system. The aim of the study was to evaluate and compare the changes of contractile properties of extensors and flexors of knee joint measured in isokinetic and isotonic regimes under conditions of simulated microgravity (7 days dry immersion).     

Human soleus fibers contractile characteristics and sarcomeric cytoskeletal proteins after gravitational unloading. Contribution of support stimulus

The effects of support withdrawal and support stimulation on the contractile characteristics of human soleus fibers and cellular factors which influence them were studied. The experimental model of the "dry" head-out water immersion was used in the study. In this model, the hydrostatic pressure on different sites of the body surface are equal so that the experimental conditions are close to the complete supportlessness. A 7-day exposure to dry immersion resulted in a decrease in the maximal isometric tension of the skinned fibers, a decline in the myofibrillar Ca2+-sensitivity, and the relative loss of the titin and nebulin content. A significant decrease in the percentage of fibers containing slow myosin heavy chains was also observed after dry immersion. The application of the mechanical stimulator influencing the plantar support zones with a pressure of 0.2 +/- 0.15 kg/cm2 6 times a day for 20 minutes of each hour brought about a complete prevention of the above listed effects of dry immersion. The data obtained allow one to conclude that the decline in maximal tension and Ca2+-sensitivity as well as myosin shift and loss of sarcomeric cytoskeletal proteins are associated with the support withdrawal during the exposure to dry immersion.     

Electrical stimulation of the posterior limb of the internal capsule for treatment of thalamic pain

Electrical stimulation of the posterior limb of the internal capsule was performed in 7 patients with thalamic pain who had developed dysesthesia, hyperpathia and/or spontaneous burning pain. Ramped bipolar stimulation elicited sensory responses, such as warm or comfortable sensation. Follow-up from 9 months to 2 years and 7 months showed that 3 patients had a good result, two had fair and the remaining two had poor results. No serious side effects were seen. The mechanism of pain relief by the internal capsule stimulation is discussed.     

Priming of macrophages with lipopolysaccharide potentiates P2X7-mediated cell death via a caspase-1-dependent mechanism, independently of cytokine production.

ATP stimulation of cell surface P2X7 receptors results in cytolysis and cell death of macrophages. Activation of this receptor in bacterial lipopolysaccharide (LPS)-activated macrophages or monocytes also stimulates processing and release of the cytokine interleukin-1beta(IL-1beta) through activation of caspase-1. The cytokine interleukin 18 (IL-18) is also cleaved by caspase-1 and shares pro-inflammatory characteristics with IL-1beta. The objective of the present study was to test the hypothesis that IL-1beta, IL-18, and/or caspase-1 activation contribute directly to macrophage cell death induced by LPS and ATP. Macrophages were cultured from normal mice or those in which genes for the P2X7 receptor, IL-1beta, IL-1alpha, IL-18, or caspase-1 had been deleted. Our data confirm the importance of the P2X7 receptor in ATP-stimulated cell death and IL-1beta release from LPS-primed macrophages. We demonstrate that prolonged stimulation with ATP leads to cell death, which is partly dependent on LPS priming and caspase-1, but independent of cytokine processing and release. We also provide evidence that LPS priming of macrophages makes them highly susceptible to the toxic effects of brief exposure to ATP, which leads to rapid cell death by a mechanism that is dependent on caspase-1 but, again, independent of cytokine processing and release.     

Interleukin (IL)-7 induces rapid activation of Pyk2, which is bound to Janus kinase 1 and IL-7Ralpha

Interleukin-7 (IL-7) receptor signaling begins with activation of the Janus tyrosine kinases Jak1 and Jak3, which are associated with the receptor complex. To identify potential targets of these kinases, we examined Pyk2 (a member of the focal adhesion kinase family) using an IL-7-dependent murine thymocyte line, D1. We demonstrate that stimulation of D1 (or normal pro-T) cells by IL-7 rapidly increased tyrosine phosphorylation and enzymatic activity of Pyk2, with kinetics slightly lagging that of Jak1 and Jak3 phosphorylation. Conversely, IL-7 withdrawal resulted in a marked decrease of Pyk2 phosphorylation. Pyk2 was found to be physically associated with Jak1 prior to IL-7 stimulation and to increase its association with IL-7Ralpha chain following IL-7 stimulation. Pyk2 appeared to be involved in cell survival, because antisense Pyk2 accelerated the cell death process. Activation of Pyk2 via the muscarinic and nicotinic receptors using carbachol or via intracellular Ca(2+) rise using ionomycin/phorbol myristate acetate promoted survival in the absence of IL-7. These data support a role for Pyk2 in coupling Jak signaling to the trophic response.     

Basal activation of the P2X7 ATP receptor elevates mitochondrial calcium and potential, increases cellular ATP levels, and promotes serum-independent growth

P2X7 is a bifunctional receptor (P2X7R) for extracellular ATP that, depending on the level of activation, forms a cation-selective channel or a large conductance nonselective pore. The P2X7R has a strong proapoptotic activity but can also support growth. Here, we describe the mechanism involved in growth stimulation. Transfection of P2X7R increases resting mitochondrial potential (delta psi(mt)), basal mitochondrial Ca2+ ([Ca2+]mt), intracellular ATP content, and confers ability to grow in the absence of serum. These changes require a full pore-forming function, because they are abolished in cells transfected with a mutated P2X7R that retains channel activity but cannot form the nonselective pore, and depend on an autocrine/paracrine tonic stimulation by secreted ATP. On the other hand, sustained stimulation of P2X7R causes a delta psi(mt) drop, a large increase in [Ca2+]mt, mitochondrial fragmentation, and cell death. These findings reveal a hitherto undescribed mechanism for growth stimulation by a plasma membrane pore.     

Modulation of the Aplysia gill withdrawal reflex by dopamine

The ability of dopamine to modulate gill contractions was tested in Aplysia. When dopamine was perfused through the gill vasculature, gill contractions caused by siphon stimulation (gill withdrawal reflex) and by depolarization of the gill motor neuron L7 were increased in amplitude, as compared with those evoked during seawater perfusion. Habituation of gill movements, brought about by repetitive stimulation of the siphon or of L7, was prevented by dopamine. Despite the absence of reflex habituation, the number of action potentials in central gill motor neurons, evoked by siphon stimulation, showed normal decrement. Dopamine's effects were blocked when the ctenidial nerve was cut or when L7 hyperpolarized. These data suggest that dopamine acts peripherally to increase the efficacy of L7's synaptic transmission onto gill muscle or elements of the gill neural plexus.     

Regional differences in mechanisms of cerebral circulatory response to neuronal activation

Vibrissal stimulation raises cerebral blood flow (CBF) in the ipsilateral spinal and principal sensory trigeminal nuclei and contralateral ventroposteromedial (VPM) thalamic nucleus and barrel cortex. To investigate possible roles of adenosine and nitric oxide (NO) in these increases, local CBF was determined during unilateral vibrissal stimulation in unanesthetized rats after adenosine receptor blockade with caffeine or NO synthase inhibition with N(G)-nitro-L-arginine methyl ester (L-NAME) or 7-nitroindazole (7-NI). Caffeine lowered baseline CBF in all structures but reduced the percent increase during stimulation only in the two trigeminal nuclei. L-NAME and 7-NI lowered baseline CBF but reduced the percent increase during stimulation only in the higher stations of this sensory pathway, i.e., L-NAME in the VPM nucleus and 7-NI in both the VPM nucleus and barrel cortex. Combinations of caffeine with 7-NI or L-NAME did not have additive effects, and none alone or in combination completely eliminated functional activation of CBF. These results suggest that caffeine-sensitive and NO-dependent mechanisms are involved but with different regional distributions, and neither fully accounts for the functional activation of CBF.     

Renin release in anesthetized rats is enhanced by the calmodulin antagonist W-7

In foregoing work, we found that the release of renin from rat kidney cortical slices was stimulated by the calmodulin antagonist W-7. The present work was done to determine whether W-7 would stimulate renin release in vivo. W-7 and its control agent, W-5 were directly infused into the renal artery of anesthetized rats. W-7 and W-5, at 50 micrograms/kg/min, produced no significant effects on renin release. Infusion of W-7 at 100 micrograms/kg/min resulted in a marked stimulation of renin release, but there was no significant alteration in the release when the same dose of W-5 was infused. Both compounds elicited a slight decrease in renal blood flow. The alterations in renin release and renal blood flow seen with W-7 were not affected by pretreatment with phentolamine or propranolol. As W-7 stimulates renin release in vivo, the hypothesis that Ca2+-calmodulin plays an inhibitory role in renin release from the kidney is given added support.     

Activation of the Transcription Factor FosB/Activating Protein-1 (AP-1) Is a Prominent Downstream Signal of the Extracellular Nucleotide Receptor P2RX7 in Monocytic and Osteoblastic Cells

Activation of the ionotropic P2RX7 nucleotide receptor by extracellular ATP has been implicated in modulating inflammatory disease progression. Continuous exposure of P2RX7 to ligand can result in apoptosis in many cell types, including monocytic cells, whereas transient activation of P2RX7 is linked to inflammatory mediator production and the promotion of cell growth. Given the rapid hydrolysis of ATP in the circulation and interstitial space, transient activation of P2RX7 appears critically important for its action, yet its effects on gene expression are unclear. The present study demonstrates that short-term stimulation of human and mouse monocytic cells as well as mouse osteoblasts with P2RX7 agonists substantially induces the expression of several activating protein-1 (AP-1) members, particularly FosB. The potent activation of FosB after P2RX7 stimulation is especially noteworthy considering that little is known concerning the role of FosB in immunological regulation. Interestingly, the magnitude of FosB activation induced by P2RX7 stimulation appears greater than that observed with other known inducers of FosB expression. In addition, we have identified a previously unrecognized role for FosB in osteoblasts with respect to nucleotide-induced expression of cyclooxygenase-2 (COX-2), which is the rate-limiting enzyme in prostaglandin biosynthesis from arachidonic acid and is critical for osteoblastic differentiation and immune behavior. The present studies are the first to link P2RX7 action to FosB/AP-1 regulation in multiple cell types, including a role in nucleotide-induced COX-2 expression, and support a role for FosB in the control of immune and osteogenic function by P2RX7.     

Effect of mechanical stimulation of the support zones of soles on the muscle stiffness in 7-day dry immersion

Stiffness of m. soleus (Sol.) and m. tibialis anterior (TA) was evaluated in 16 volunteers during exposure to 7-days dry immersion alone and to the combination of immersion and mechanic stimulation of foot support zones. It was shown that Sol. stiffness decreased progressively starting from day-1 of immersion, whereas TA stiffness, on the contrary, made a sharp rise. Mechanic stimulation of foot support zones slowed down the rate and extent of changes in both muscles.     

Leukocyte complement: a possible role for C5 in lymphocyte stimulation

The results presented here show that Fab' antibody fragments directed to complement proteins C5, C6, and C7 inhibit lymphocyte stimulation in mixed lymphocyte culture (MLC) by up to 65%, as determined by decreased incorporation of 3H-thymidine. Lymphocyte stimulation induced by PHA-mitogen was also inhibited up to 100% by anti-C5 Fab'. Specificity of these reactions was established by the findings that goat anti-C5 or murine hybridoma anti-C5 both inhibited MLC; the inhibitory activity of anti-C5 Fab' was absorbed with highly purified C5 (but not with C3), and antibody directed to C3 did not inhibit lymphocyte stimulation by MLC or PHA. The effects of anti-C5 were exerted in a nontoxic manner. Cleavage of lymphocyte associated C5 with factor B (Bb) or with trypsin resulted in stimulation of lymphocyte thymidine incorporation. Purified C5a was found to induce lymphocyte stimulation in serum-free medium in pulse-chase types of experiments. Anti-C6 and C7 Fab' also inhibited lymphocyte stimulation induced in one-way MLC. These results suggest that C5, C5a, and/or C6 and C7 may play a role in triggering of lymphocyte blastogenesis.     

Serum creatine kinase levels and the number of sarcolemmal dystrophin disruptions in human skeletal muscle fibers under conditions of 7-day "dry" immersion

9 male volunteers took part in the experiment. They were divided in two groups. 5 volunteers (control group) have been in "dry" immersion for 7 days. 4 volunteers (stimulated group) in addition to "dry" immersion were treated with artificial support stimulation. We investigated the number of muscle fibers with the disruptions of sarcolemmal dystrophin and serum creatine kinase levels. 7-day "dry" immersion does not change the mean number of muscle fibers with dystrophin disruptions, it leads to significant decrease of serum creatine kinase levels and does not influence on the sensitivity of sarcolemma to injury. Artificial support stimulation does not influence on these parameters.     

Effects of intraoral mechanoreceptor stimulation on reticular formation neurones, in the rabbit

The effects of intraoral mechanoreceptor stimulation on the firing rate of single neurones of the brain stem reticular formation (RF) were investigated in rabbits. 30% of RF neurones responded to periodontal mechanoreceptor stimulation; 16% to mucosal mechanoreceptor stimulation and 6% to both types of stimuli. Periodontal stimulation induced mainly inhibitory effects localized within the mesencephalic and rostral pontine RF. Among periodontal afferents incisors were the most widely represented. The effects of mucosal mechanoreceptor stimulation were predominant in the medullary and pontine RF and they were mainly excitatory. The present results support the hypothesis that brain stem RF neurones can be recruited into regulating mastication and biting also by stimulation of intraoral mechanoreceptors.     

Medullary substrate and differential cardiovascular responses during stimulation of specific acupoints

Electroacupuncture (EA) at P5-P6 acupoints overlying the median nerve reduces premotor sympathetic cardiovascular neuronal activity in the rostral ventral lateral medulla (rVLM) and visceral reflex pressor responses. In previous studies, we have noted different durations of influence of EA comparing P5-P6 and S36-S37 acupoints, suggesting that point specificity may exist. The purpose of this study was to evaluate the influence of stimulating P5-P6 (overlying the median nerve), LI4-L7 (overlying branches of the median nerve and the superficial radial nerve), LI6-LI7 (overlying the superficial radial nerve), LI10-LI11 (overlying the deep radial nerves), S36-S37 (overlying the deep peroneal nerves), or K1-B67 (overlying terminal branches of the tibial nerves) specific acupoints, overlying deep and superficial somatic nerves, on the excitatory cardiovascular reflex and rVLM responses evoked by stimulation of chemosensitive receptors in the cat's gallbladder with bradykinin (BK) or direct splanchnic nerve (SN) stimulation. We observed point-specific differences in magnitude and duration of EA inhibition between P5-P6 or LI10-LI11 and LI4-L7 or S36-S37 in responses to 30-min stimulation with low-frequency, low-current EA. EA at LI6-LI7 and K1-B67 acupoints as well as direct stimulation of the superficial radial nerve did not cause any cardiovascular or rVLM neuronal effects. Cardiovascular neurons in the rVLM, a subset of which were classified as premotor sympathetic cells, responded to brief (30 s) stimulation of the SN as well as acupoints P5-P6, LI10-LI11, LI4-L7, S36-S37, LI6-LI7, or K1-B67, or underlying somatic pathways in a fashion similar to the reflex responses. In fact, we observed a significant linear relationship (r(2) = 0.71) between the evoked rVLM response and reflex change in mean arterial blood pressure. In addition, EA stimulation at P5-P6 and LI4-L7 decreased rVLM neuronal activity by 41 and 12%, respectively, for >1 h, demonstrating that prolonged input into the medulla during stimulation of somatic nerves, depending on the degree of convergence, leads to more or less inhibition of activity of these cardiovascular neurons. Thus EA at acupoints overlying deep and superficial somatic nerves leads to point-specific effects on cardiovascular reflex responses. In a similar manner, sympathetic cardiovascular rVLM neurons that respond to both visceral (reflex) and somatic (EA) nerve stimulation manifest graded responses during stimulation of specific acupoints, suggesting that this medullary region plays a role in site-specific inhibition of cardiovascular reflex responses by acupuncture.