Calcium and Vitamin D increase mRNA levels for the growth control hIK1 channel in human epidermal keratinocytes but functional channels are not observed

BACKGROUND: Intermediate-conductance, calcium-activated potassium channels (IKs) modulate proliferation and differentiation in mesodermal cells by enhancing calcium influx, and they contribute to the physiology of fluid movement in certain epithelia. Previous reports suggest that IK channels stimulate proliferative growth in a keratinocyte cell line; however, because these channels indirectly promote calcium influx, a critically unique component of the keratinocyte differentiation program, an alternative hypothesis is that they would be anti-proliferative and pro-differentiating. This study addresses these hypotheses. METHODS: Real-time PCR, patch clamp electrophysiology, and proliferation assays were used to determine if human IK1 (hIK1) expression and function are correlated with either proliferation or differentiation in cultured human skin epidermal keratinocytes, and skin biopsies grown in explant culture. RESULTS: hIK1 mRNA expression in human keratinocytes and skin was increased in response to anti-proliferative/pro-differentiating stimuli (elevated calcium and Vitamin D). Correspondingly, the hIK1 agonist 1-EBIO inhibited keratinocyte proliferation suggesting that the channel could be anti-proliferative and pro-differentiating. However, this proliferative inhibition by 1-EBIO was not reversed by a panel of hIK1 blockers, calling into question the mechanism of 1-EBIO action. Subsequent patch clamp electrophysiological analysis failed to detect hIK1 channel currents in keratinocytes, even those expressing substantial hIK1 mRNA in response to calcium and Vitamin D induced differentiation. Identical electrophysiological recording conditions were then used to observe robust IK1 currents in fibroblasts which express IK1 mRNA levels comparable to those of keratinocytes. Thus, the absence of observable hIK1 currents in keratinocytes was not a function of the electrophysiological techniques. CONCLUSION: Human keratinocyte differentiation is stimulated by calcium mobilization and influx, and differentiation stimuli coordinately upregulate mRNA levels of the calcium-activated hIK1 channel. This upregulation is paradoxical in that functional hIK1 channels are not observed in cultured keratinocytes. It appears, therefore, that hIK1 does not contribute to the functional electrophysiology of primary human keratinocytes, nor intact human skin. Further, the results indicate caution is required when interpreting experiments utilizing pharmacological hIK1 modulators in human keratinocytes

Effect of sedation and analgesia on postoperative amplitude-integrated EEG in newborn cardiac patients

The aim of this study is to describe the effect of sedation and analgesia on postoperative amplitude-integrated EEG (aEEG) in newborns with congenital heart disease (CHD) undergoing heart surgery. This is a consecutive series of 26 newborns with CHD of which 16 patients underwent cardiopulmonary bypass (CPB) surgery and 10 patients did not. aEEG was monitored for at least 12 h preoperatively and started within the first 6 h postoperatively for 48 h. Outcome was assessed at 1 year of age. All 26 patients showed a normal preoperative continuous cerebral activity with sleep-wake cycles (SWC). The postoperative duration to return to normal background activity with SWC was similar for both groups. Independent of group assignment, patients requiring midazolam had a significantly later onset of a normal SWC than those without midazolam (p=0.03). Three patients in the CPB group and two in the non-CPB group showed continuous low voltage or flat trace after administration of fentanyl. These changes did not correlate with neurodevelopmental outcome. Sedation with midazolam has a transient effect on the background activity, whereas fentanyl can induce a severe pathologic background pattern. The significance of these changes on outcome is not yet clear. Thus, more attention should be paid to these effects when interpreting aEEG in this population