COVID-19 lockdown and the rate of central precocious puberty.

PURPOSE: The aim of our study was to compare the incidence of idiopathic central precocious puberty (CPP) in our highly specialized Endocrinological Center before and after the onset of COVID-19 lockdown; we also aimed to identify any potential difference between girls with CPP from the two different time periods. METHODS: We retrospectively analyzed the auxological profile of 49 girls with idiopathic CPP: 30 with pre-lockdown onset and 19 with post-lockdown onset of the disease. We collected patients' characteristics (medical history, physical examination, baseline and dynamic hormonal assessment, bone age, pelvic ultrasound) and compared them between the two groups. RESULTS: We registered an almost threefold increase in CPP incidence in the 2020-2021 period compared to the previous six years. In post-lockdown patients we found a trend for an earlier diagnosis in terms of both chronological age (p 0.0866) and days between the onset of first pubertal signs and diagnosis (p 0.0618). We also found that post-lockdown patients had a significantly lower hypothalamus-pituitary-gonadal axis activation (lower ∆LH% after GnRH test, p 0.0497), a significantly lower increase in bone age calculated at RUS with TW3 method (p 0.0438) and a significantly reduced ovarian activation in females (lower delta-4-androstenedione levels, p 0.0115). Interestingly, post-lockdown patients were born from mothers with an older age at menarche (p 0.0039). CONCLUSIONS: Besides confirming a significant increase in new diagnoses of CPP in the post-lockdown period, our findings among Post-lockdown girls also suggest a less progressive form of CPP and a stronger environmental influence compared to genetic background in determining the timing of pubertal onset.

Body weight variation is not an independent factor in the determination of functional hypothalamic amenorrhea in anorexia nervosa.

OBJECTIVE: Functional hypothalamic amenorrhea (FHA) is one of the foremost manifestations in anorexia nervosa (AN), but a subset of patients have menses despite marked weight loss and underweight. The aim of our study was to investigate parameters potentially influencing FHA in AN. DESIGN AND METHODS: In this observational retrospective study, we selected 114 female patients with AN who completed a 12 months semi-residential rehabilitation program and a subsequent 12 months outpatient follow-up. We divided our sample into three groups: "Group 0" patients who experienced FHA and recovered their menses, "Group 1" persistent FHA, "Group 2" never experienced FHA, and looked for clinical and hormonal correlations. RESULTS: At the enrollment, the BMI was higher in Group 2 than in Group 1 (p = 0.0202), but the last follow-up weight was higher in Group 1 (p < 0.0001) despite persistent amenorrhea. At logistic regression, the higher BMI at which patients experienced amenorrhea was the main prediction factor for persistent FHA. Notwithstanding comparable leptin levels at admission, they improved significantly at discharge only in Groups 0 and 2 (p = 0.0054 and p = 0.0104, respectively). FT3 at admission was significantly higher in Group 2 than in Group 0 (p = 0.0249). CONCLUSIONS: FHA does not correlate strictly with body weight variations in AN patients, indicating a multifactorial origin, likely including an individual predisposition. Higher FT3 levels identify patients who continue having menses at extremely low BMI. AN patients with persistent FHA constitute a subgroup in whom estroprogestins should be considered after significant weight recovery to prevent prolonged tissue hypoestrogenism.

Stochastic epigenetic mutations as possible explanation for phenotypical discordance among twins with congenital hypothyroidism.

PURPOSE: The elevated frequency of discordance for congenital hypothyroidism (CH) phenotype between monozygotic twins suggests the involvement of non-mendelian mechanisms. The aim of the study was to investigate the role of epigenetics in CH pathogenesis. METHODS: A genome-wide DNA methylation analysis was performed on the peripheral blood of 23 twin pairs (10 monozygotic and 13 dizygotic), 4 concordant and 19 discordant pairs for CH at birth. RESULTS: Differential methylation analysis did not show significant differences in methylation levels between CH cases and controls, but a different methylation status of several genes may explain the CH discordance of a monozygotic twin couple carrying a monoallelic nonsense mutation of DUOX2. In addition, the median number of hypo-methylated Stochastic Epigenetic Mutations (SEMs) resulted significantly increased in cases compared to controls. The prioritization analysis for CH performed on the genes epimutated exclusively in the cases identified SLC26A4, FOXI1, NKX2-5 and TSHB as the genes with the highest score. The analysis of significantly SEMs-enriched regions led to the identification of two genes (FAM50B and MEG8) that resulted epigenetically dysregulated in cases. CONCLUSION: Epigenetic modifications may potentially account for CH pathogenesis and explain discordance among monozygotic twins.

A systematic review on drugs for synaptic plasticity in the treatment of dementia.

The aim of the present systematic review (SR) was to provide an overview of all published and unpublished clinical trials investigating the safety and efficacy of disease-modifying drugs targeting synaptic plasticity in dementia. Searches on CT.gov and EuCT identified 27 trials (4 phase-1, 1 phase-1/2, 18 phase-2, 1 phase-2/3, 1 phase-3, 1 phase-4, and 1 not reported). Twenty of them completed, and seven are currently active or enrolling. The structured bibliographic searches yielded 3585 records. A total of 12 studies were selected on Levetiracetam, Masitinib, Saracatinib, BI 40930, Bryostatin 1, PF-04447943 and Edonerpic drugs. We used RoB tool for quality analysis of randomized studies. Efficacy was assessed as a primary outcome in all studies except one and the main scale used was ADAS-Cog (7 studies), MMSE and CDR (4 studies). Safety and tolerability were reported in eleven studies. The incidence of SAEs was similar between treatment and placebo. At the moment, only one molecule reached phase-3. This could suggest that research on these drugs is still preliminary. Of all, three studies reported promising results on Levetiracetam, Bryostatin 1 and Masitinib.

Development and validation of circulating tumour cell enumeration (Epic Sciences) as a prognostic biomarker in men with metastatic castration-resistant prostate cancer.

PURPOSE: To evaluate the prognostic significance of circulating tumour cell (CTC) number determined on the Epic Sciences platform in men with metastatic castration-resistant prostate cancer (mCRPC) treated with an androgen receptor signalling inhibitor (ARSI). PATIENTS AND METHODS: A pre-treatment blood sample was collected from men with progressing mCRPC starting either abiraterone or enzalutamide as a first-, second- or third-line systemic therapy at Memorial Sloan Kettering Cancer Center (Discovery cohort, N = 171) or as a first- or second-line therapy as part of the multicenter PROPHECY trial (NCT02269982) (Validation cohort, N = 107). The measured CTC number was then associated with overall survival (OS) in the Discovery cohort, and progression-free survival (PFS) and OS in the Validation cohort. CTC enumeration was also performed on a concurrently obtained blood sample using the CellSearch® Circulating Tumor Cell Kit. RESULTS: In the MSKCC Discovery cohort, CTC count was a statistically significant prognostic factor of OS as a dichotomous (<3 CTCs/mL versus ≥ 3 CTCs/mL; hazard ratio [HR] = 1.8 [95% confidence interval {CI} 1.3-3.0]) and a continuous variable when adjusting for line of therapy, presence of visceral metastases, prostate-specific antigen, lactate dehydrogenase and alkaline phosphatase. The findings were validated in an independent datas et from PROPHECY (HR [95% CI] = 1.8 [1.1-3.0] for OS and 1.7 [1.1-2.9] for PFS). A strong correlation was also observed between CTC counts determined in matched samples on the CellSearch® and Epic platforms (r = 0.84). CONCLUSION: The findings validate the prognostic significance of pretreatment CTC number determined on the Epic Sciences platform for predicting OS in men with progressing mCRPC starting an ARSI.

Research participation among state and local public health emergency preparedness and response programs.

OBJECTIVES: The objective of our study was to assess whether state and local health staff participated in public health emergency preparedness research activities and what partner organizations they collaborated with on research. STUDY DESIGN: This is a cross-sectional study. METHODS: Data were derived from a 2014 web-based survey of state, territorial, and local health departments conducted by the Centers for Disease Control and Prevention and NORC at the University of Chicago as part of a larger project to assess the public health emergency preparedness and response research priorities of state and local health departments. RESULTS: Overall, 30% of survey respondents indicated that health department staff were involved in public health preparedness and response research-related activities. Thirty-four percent indicated that they were extremely or moderately familiar with emergency preparedness research and literature. Approximately 67% of respondents reported interest in receiving additional information and/or training related to the preparedness research and literature. The most frequently reported partners for collaboration in preparedness research-related activities were schools of public health (34%). CONCLUSIONS: Our findings suggest that there is health department interest in learning more about preparedness and response science and that additional efforts are needed to increase health department participation in public health emergency preparedness and response research-related activities.

Nanodiamonds-induced effects on neuronal firing of mouse hippocampal microcircuits.

Fluorescent nanodiamonds (FND) are carbon-based nanomaterials that can efficiently incorporate optically active photoluminescent centers such as the nitrogen-vacancy complex, thus making them promising candidates as optical biolabels and drug-delivery agents. FNDs exhibit bright fluorescence without photobleaching combined with high uptake rate and low cytotoxicity. Focusing on FNDs interference with neuronal function, here we examined their effect on cultured hippocampal neurons, monitoring the whole network development as well as the electrophysiological properties of single neurons. We observed that FNDs drastically decreased the frequency of inhibitory (from 1.81 Hz to 0.86 Hz) and excitatory (from 1.61 to 0.68 Hz) miniature postsynaptic currents, and consistently reduced action potential (AP) firing frequency (by 36%), as measured by microelectrode arrays. On the contrary, bursts synchronization was preserved, as well as the amplitude of spontaneous inhibitory and excitatory events. Current-clamp recordings revealed that the ratio of neurons responding with AP trains of high-frequency (fast-spiking) versus neurons responding with trains of low-frequency (slow-spiking) was unaltered, suggesting that FNDs exerted a comparable action on neuronal subpopulations. At the single cell level, rapid onset of the somatic AP ("kink") was drastically reduced in FND-treated neurons, suggesting a reduced contribution of axonal and dendritic components while preserving neuronal excitability.

Knock-down of synapsin alters cell excitability and action potential waveform by potentiating BK and voltage-gated Ca(2+) currents in Helix serotonergic neurons.

Synapsins (Syns) are an evolutionarily conserved family of presynaptic proteins crucial for the fine-tuning of synaptic function. A large amount of experimental evidences has shown that Syns are involved in the development of epileptic phenotypes and several mutations in Syn genes have been associated with epilepsy in humans and animal models. Syn mutations induce alterations in circuitry and neurotransmitter release, differentially affecting excitatory and inhibitory synapses, thus causing an excitation/inhibition imbalance in network excitability toward hyperexcitability that may be a determinant with regard to the development of epilepsy. Another approach to investigate epileptogenic mechanisms is to understand how silencing Syn affects the cellular behavior of single neurons and is associated with the hyperexcitable phenotypes observed in epilepsy. Here, we examined the functional effects of antisense-RNA inhibition of Syn expression on individually identified and isolated serotonergic cells of the Helix land snail. We found that Helix synapsin silencing increases cell excitability characterized by a slightly depolarized resting membrane potential, decreases the rheobase, reduces the threshold for action potential (AP) firing and increases the mean and instantaneous firing rates, with respect to control cells. The observed increase of Ca(2+) and BK currents in Syn-silenced cells seems to be related to changes in the shape of the AP waveform. These currents sustain the faster spiking in Syn-deficient cells by increasing the after hyperpolarization and limiting the Na(+) and Ca(2+) channel inactivation during repetitive firing. This in turn speeds up the depolarization phase by reaching the AP threshold faster. Our results provide evidence that Syn silencing increases intrinsic cell excitability associated with increased Ca(2+) and Ca(2+)-dependent BK currents in the absence of excitatory or inhibitory inputs.

Perceived psychosocial stress and glucose intolerance among pregnant Hispanic women.

AIM: Prior literature suggests a positive association between psychosocial stress and the risk of diabetes in non-pregnant populations, but studies during pregnancy are sparse. We evaluated the relationship between stress and glucose intolerance among 1115 Hispanic (predominantly Puerto Rican) prenatal care patients in Proyecto Buena Salud, a prospective cohort study in Western Massachusetts (2006-2011). METHODS: Cohen's Perceived Stress Scale (PSS-14) was administered in early (mean = 12.3 weeks gestation; range 4.1-18 weeks) and mid- (mean = 21.3 weeks gestation; range 18.1-26 weeks) pregnancy. Participants were classified as having a pregnancy complicated by gestational diabetes mellitus, impaired glucose tolerance, and abnormal glucose tolerance, based on the degree of abnormality on glucose tolerance testing between 24 and 28 weeks of gestation. RESULTS: The prevalence of gestational diabetes mellitus, impaired glucose tolerance, and abnormal glucose tolerance was 4.1%, 7.2%, and 14.5%, respectively. Absolute levels of early or mid-pregnancy stress were not significantly associated with glucose intolerance. However, participants with an increase in stress from early to mid-pregnancy had a 2.6-fold increased odds of gestational diabetes mellitus (95% confidence intervals: 1.0-6.9) as compared to those with no change or a decrease in stress after adjusting for age and pre-pregnancy body mass index. In addition, every one-point increase in stress scores was associated with a 5.5mg/dL increase in screening glucose level (ß=5.5; standard deviation=2.8; P=0.05), after adjusting for the same variables. CONCLUSION: In this population of predominantly Puerto Rican women, stress patterns during pregnancy may influence the risk of glucose intolerance.

Methicillin-resistant Staphylococcus aureus ST30-SCCmec IVc clone as the major cause of community-acquired invasive infections in Argentina.

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have become a major concern worldwide. We conducted a prospective multicenter study of invasive CA-MRSA to evaluate clinical features and genotype of strains causing invasive infections in Argentina. A total of 55 patients with invasive CA-MRSA infections were included. Most patients (60%) had bloodstream infections, 42% required admission to intensive care unit and 16% died. No CA-MRSA isolates were multiresistant (resistant ⩾3 classes of antibiotics). All isolates carried Panton-Valentine leukocidin (PVL) genes and staphylococcal cassette chromosome (SCCmec) type IV. The majority CA-MRSA strains belonged to ST30 and had identical pulsed-field gel electrophoresis (PFGE) patterns, qualifying as a clonal dissemination of a highly transmissible strain. The main clone recovered from patients with CA-MRSA invasive infections was genotyped as pulsed-field gel electrophoresis type C-ST30, SCCmec type IVc-spa type 019, PVL positive. It has become predominant and replaced the previously described CA-MRSA clone (PFGE type A, ST5, SCCmec type IV, spa type 311).

Cav1.3 and Cav1.2 channels of adrenal chromaffin cells: emerging views on cAMP/cGMP-mediated phosphorylation and role in pacemaking.

Voltage-gated Ca²âº channels (VGCCs) are voltage sensors that convert membrane depolarizations into Ca²âº signals. In the chromaffin cells of the adrenal medulla, the Ca²âº signals driven by VGCCs regulate catecholamine secretion, vesicle retrievals, action potential shape and firing frequency. Among the VGCC-types expressed in these cells (N-, L-, P/Q-, R- and T-types), the two L-type isoforms, Ca(v)1.2 and Ca(v)1.3, control key activities due to their particular activation-inactivation gating and high-density of expression in rodents and humans. The two isoforms are also effectively modulated by G protein-coupled receptor pathways delimited in membrane micro-domains and by the cAMP/PKA and NO/cGMP/PKG phosphorylation pathways which induce prominent Ca²âº current changes if opposingly regulated. The two L-type isoforms shape the action potential and directly participate to vesicle exocytosis and endocytosis. The low-threshold of activation and slow rate of inactivation of Ca(v)1.3 confer to this channel the unique property of carrying sufficient inward current at subthreshold potentials able to activate BK and SK channels which set the resting potential, the action potential shape, the cell firing mode and the degree of spike frequency adaptation during spontaneous firing or sustained depolarizations. These properties help chromaffin cells to optimally adapt when switching from normal to stress-mimicking conditions. Here, we will review past and recent findings on cAMP- and cGMP-mediated modulations of Ca(v)1.2 and Ca(v)1.3 and the role that these channels play in the control of chromaffin cell firing. This article is part of a Special Issue entitled: Calcium channels.

Calcium channel types contributing to chromaffin cell excitability, exocytosis and endocytosis.

Voltage gated Ca(2+) channels are effective voltage sensors of plasma membrane which convert cell depolarizations into Ca(2+) signaling. The chromaffin cells of the adrenal medulla utilize a large number of Ca(2+) channel types to drive the Ca(2+)-dependent release of catecholamines into blood circulation, during normal or stress-induced conditions. Some of the Ca(2+) channels expressed in chromaffin cells (L, N, P/Q, R and T), however, do not control only vesicle fusion and catecholamine release. They also subserve a variety of key activities which are vital for the physiological and pathological functioning of the cell, like: (i) shaping the action potentials of electrical oscillations driven either spontaneously or by ACh stimulation, (ii) controlling the action potential frequency of tonic or bursts firing, (iii) regulating the compensatory and excess endocytosis following robust exocytosis and (iv) driving the remodeling of Ca(2+) signaling which occurs during stressors stimulation. Here, we will briefly review the well-established properties of voltage-gated Ca(2+) channels accumulated over the past three decades focusing on the most recent discoveries on the role that L- (Cav1.2, Cav1.3) and T-type (Cav3.2) channels play in the control of excitability, exocytosis and endocytosis of chromaffin cells in normal and stress-mimicking conditions.

Nanocrystalline diamond microelectrode arrays fabricated on sapphire technology for high-time resolution of quantal catecholamine secretion from chromaffin cells.

The quantal release of oxidizable molecules can be successfully monitored by means of polarized carbon fiber microelectrodes (CFEs) positioned in close proximity to the cell membrane. To partially overcome certain CFE limitations, mainly related to their low spatial resolution and lack of optical transparency, we developed a planar boron-doped nanocrystalline diamond (NCD) prototype, grown on a transparent sapphire wafer. Responsiveness to applied catecholamines as well as the electrochemical and optical properties of the NCD-based device were first characterized by cyclic voltammetry and optical transmittance measurements. By stimulating chromaffin cells positioned on the device with external KCl, well-resolved quantal exocytotic events could be detected either from one NCD microelectrode, or simultaneously from an array of four microelectrodes, indicating that the chip is able to monitor secretory events (amperometric spikes) from a number of isolated chromaffin cells. Spikes detected by the planar NCD device had comparable amplitudes, kinetics and vesicle diameter distributions as those measured by conventional CFEs from the same chromaffin cell.

PDE type-4 inhibition increases L-type Ca(2+) currents, action potential firing, and quantal size of exocytosis in mouse chromaffin cells.

We studied the effects of the cAMP-hydrolyzing enzyme phosphodiesterase type-4 (PDE4) on the L-type Ca(2+) channels (LTCCs) and Ca(2+)-dependent secretion in mouse chromaffin cells (MCCs). The selective PDE4 inhibitor rolipram (3 microM) had a specific potentiating action on Ca(2+) currents of MCCs (40% increase within 3 min). A similar effect was produced by the selective beta(1)-AR agonist denopamine (1 microM) and by the unselective PDEs inhibitor IBMX (100 microM). Rolipram and denopamine actions were selective for LTCCs, and the Ca(2+) current increase remained unchanged if the two compounds were applied simultaneously. This suggests that at rest, LTCCs in MCCs are down-regulated by the low levels of cAMP determined by PDE4 activity and that LTCCs can be up-regulated by either inhibiting PDE4 or activating beta(1)-AR. No other PDEs are likely involved in this specific action. PDE4 inhibition had also a marked effect on the spontaneous firing of resting MCCs and catecholamine secretion. Rolipram up-regulated the LTCCs contributing to the "pace-maker" current underlying action potential (AP) discharges and accelerated the firing rate, with no significant effects on AP waveform. Acceleration of AP firing was also induced by the LTCC-agonist Bay K (1 microM), while nifedipine (3 microM) reduced the firing frequency, suggesting that LTCCs and intracellular cAMP play a key role in setting the pace-maker current regulating MCCs excitability. Rolipram increased also the size of the ready-releasable pool and the quantal content of secretory vesicles without affecting their probability of release. Thus, rolipram acts on MCCs by up-regulating both exocytosis and AP firings. These two processes are effectively down-regulated by PDE4 at rest and can dramatically increase the quantity of released catecholamines when PDE4 is inhibited and/or cAMP is raised.

Allosteric modulation of alpha 7 nicotinic receptors selectively depolarizes hippocampal interneurons, enhancing spontaneous GABAergic transmission.

The role of postsynaptic nicotinic receptors for acetylcholine (nAChRs) in mediating fast neurotransmission processes in the CNS is controversial. Here we have studied the modulation of synaptic transmission by an agonist (choline) and an allosteric modulator (5-OH-indole) of alpha7 nAChRs in rat hippocampal neuronal cultures. Choline evoked a fast inactivating inward current, causing neuron depolarization and action potential discharge, thereby enhancing the spontaneous postsynaptic current activity (sPSCs). This effect was markedly enhanced when both choline and 5-OH-indole were applied together and was blocked by the selective alpha7 nAChR antagonist methyllycaconitine. This choline action was suppressed by the GABA(A) receptor antagonist bicuculline, while the glutamatergic receptor antagonist kynurenic acid had no effect. Frequency, but not amplitude or area, of both excitatory and inhibitory miniature postsynaptic currents (mEPSCs and mIPSCs) were drastically reduced when Ca(2+) influx was blocked by Cd(2+). Additionally, nAChR activation did not modify the mIPSCs. These data suggest that Ca(2+) influx through the highly Ca(2+)-permeablealpha7 nAChRs was insufficient to directly activate neurotransmitter release, suggesting that a tight colocalization of this receptor with secretory hot spots is unlikely. In a few cases, the activation of alpha7 AChRs led to a suppression of spontaneous synaptic transmission. This effect may be related to the potentiation of GABAergic interneurons that inhibit the spontaneous activity of neurons making synapses with the cell under study. We suggest that GABA release is modulated by alpha7 nAChRs. Thus, selective allosteric modulators of alpha7 nAChRs could have potential therapeutic applications in brain disorders such as epilepsy and schizophrenia and in alterations of cognition and sensory processing.

Calcium channels in chromaffin cells: focus on L and T types.

Voltage-gated Ca2+ channels (Cav) are highly expressed in the adrenal chromaffin cells of mammalian species. Besides shaping action potential waveforms, they are directly involved in the excitation-secretion coupling underlying catecholamine release and, possibly, control other Ca2+-dependent events that originate near the membrane. These functions are shared by a number of Cav channel types (L, N, P/Q, R and T) which have different structure-function characteristics and whose degree of expression changes remarkably among mammalian species. Understanding precisely the functioning of each voltage-gated Ca2+ channels is a crucial task that helps clarifying the Ca2+-dependent mechanisms controlling exocytosis during physiological and pathological conditions. In this paper, we focus on classical and new roles that L- and T-type channels play in the control of chromaffin cell excitability and neurotransmitter release. Interestingly, L-type channels are shown to be implicated in the spontaneous autorhythmicity of chromaffin cells, while T-type channels, which are absent in adult chromaffin cells, are coupled with secretion and can be recruited following long-term beta-adrenergic stimulation or chronic hypoxia. This suggests that like other cells, adrenal chromaffin cells undergo effective remodelling of membrane ion channels and cell functioning during prolonged stress conditions.

Chronic hypoxia up-regulates alpha1H T-type channels and low-threshold catecholamine secretion in rat chromaffin cells.

alpha(1H) T-type channels recruited by beta(1)-adrenergic stimulation in rat chromaffin cells (RCCs) are coupled to fast exocytosis with the same Ca(2+) dependence of high-threshold Ca(2+) channels. Here we show that RCCs exposed to chronic hypoxia (CH) for 12-18 h in 3% O(2) express comparable densities of functional T-type channels that depolarize the resting cells and contribute to low-voltage exocytosis. Following chronic hypoxia, most RCCs exhibited T-type Ca(2+) channels already available at -50 mV with the same gating, pharmacological and molecular features as the alpha(1H) isoform. Chronic hypoxia had no effects on cell size and high-threshold Ca(2+) current density and was mimicked by overnight incubation with the iron-chelating agent desferrioxamine (DFX), suggesting the involvement of hypoxia-inducible factors (HIFs). T-type channel recruitment occurred independently of PKA activation and the presence of extracellular Ca(2+). Hypoxia-recruited T-type channels were partially open at rest (T-type 'window-current') and contributed to raising the resting potential to more positive values. Their block by 50 microm Ni(2+) caused a 5-8 mV hyperpolarization. The secretory response associated with T-type channels could be detected following mild cell depolarizations, either by capacitance increases induced by step depolarizations or by amperometric current spikes induced by increased [KCl]. In the latter case, exocytotic bursts could be evoked even with 2-4 mm KCl and spike frequency was drastically reduced by 50 microm Ni(2+). Chronic hypoxia did not alter the shape of spikes, suggesting that hypoxia-recruited T-type channels increase the number of secreted vesicles at low voltages, without altering the mechanism of catecholamine release and the quantal content of released molecules.

L-type calcium channels in adrenal chromaffin cells: role in pace-making and secretion.

Voltage-gated L-type (Cav1.2 and Cav1.3) channels are widely expressed in cardiovascular tissues and represent the critical drug-target for the treatment of several cardiovascular diseases. The two isoforms are also abundantly expressed in neuronal and neuroendocrine tissues. In the brain, Cav1.2 and Cav1.3 channels control synaptic plasticity, somatic activity, neuronal differentiation and brain aging. In neuroendocrine cells, they are involved in the genesis of action potential generation, bursting activity and hormone secretion. Recent studies have shown that Cav1.2 and Cav1.3 are also expressed in chromaffin cells but their functional role has not yet been identified despite that L-type channels possess interesting characteristics, which confer them an important role in the control of catecholamine secretion during action potentials stimulation. In intact rat adrenal glands L-type channels are responsible for adrenaline and noradrenaline release following splanchnic nerve stimulation or nicotinic receptor activation. L-type channels can be either up- or down-modulated by membrane autoreceptors following distinct second messenger pathways. L-type channels are tightly coupled to BK channels and activate at relatively low-voltages. In this way they contribute to the action potential hyperpolarization and to the pace-maker current controlling action potential firings. L-type channels are shown also to regulate the fast secretion of the immediate readily releasable pool of vesicles with the same Ca(2+)-efficiency of other voltage-gated Ca(2+) channels. In mouse adrenal slices, repeated action potential-like stimulations drive L-type channels to a state of enhanced stimulus-secretion efficiency regulated by beta-adrenergic receptors. Here we will review all these novel findings and discuss the possible implication for a specific role of L-type channels in the control of chromaffin cells activity.

Fast exocytosis mediated by T- and L-type channels in chromaffin cells: distinct voltage-dependence but similar Ca2+ -dependence.

Expression, spatial distribution and specific roles of different Ca(2+) channels in stimulus-secretion coupling of chromaffin cells are intriguing issues still open to discussion. Most of the evidence supports a role of high-voltage activated (HVA) Ca(2+) channels (L-, N-, P/Q- and R-types) in the control of exocytosis: some suggesting a preferential coupling of specific Ca(2+) channel subunits with the secretory apparatus, others favoring the idea of a contribution to secretion proportional to the expression density and gating properties of Ca(2+) channels. In this work we review recent findings and bring new evidence in favor of the hypothesis that also the LVA (low-voltage-activated, T-type) Ca(2+) channels effectively control fast exocytosis near resting potential in adrenal chromaffin cells of adult rats. T-type channels recruited after long-term treatments with pCPT-cAMP (or chronic hypoxia) are shown to control exocytosis with the same efficacy of L-type channels, which are the dominant Ca(2+) channel types expressed in rodent chromaffin cells. A rigorous comparison of T- and L-type channel properties shows that, although operating at different potentials and with different voltage-sensitivity, the two channels possess otherwise similar Ca(2+)-dependence of exocytosis, size and kinetics of depletion of the immediately releasable pool and mobilize vesicles of the same quantal size. Thus, T- and L-type channels are coupled with the same Ca(2+)-efficiency to the secretory apparatus and deplete the same number of vesicles ready for release. The major difference of the secretory signals controlled by the two channels appear to be the voltage range of operation, suggesting the idea that stressful conditions (hypoxia and persistent beta-adrenergic stimulation) can lower the threshold of cell excitability by recruiting new Ca(2+) channels and activate an additional source of catecholamine secretion.

A new role for T-type channels in fast "low-threshold" exocytosis.

Evidence is accumulating on a key role of T-type channels in neurotransmitter release. Recent works have brought undisputable proofs that T-type channels are capable of controlling hormone and neurotransmitters release in association with exocytosis of large dense-core and synaptic vesicles. T-type channel-secretion coupling is not as ubiquitous as that shown for N- and P/Q-type channels in central neurons. In this case, the high-density of Cav2 channel types and co-localization to the release sites ensure high rates of vesicle release and synchronous synaptic responses. Nevertheless, when sufficiently expressed in distal dendrites and neurosecretory cells, T-type channels are able to drive the fast fusion of vesicles ready for release during "low-threshold" Ca2+-entry. T-type channels appear effectively coupled to fast vesicle depletion and may possibly regulate other Ca2+-dependent processes like vesicle recycling and vesicle mobilization from a reserve pool that are important mechanisms controlling synaptic activity during sustained stimulation. Here, we will briefly review the main findings that assign a specific task to T-type channels in fast exocytosis discussing their possible involvement in the control of the Ca2+-dependent processes regulating synaptic activity and vesicular hormone release.