Small-scale farmer responses to the double exposure of climate change and market integration.

Anthropologists have long studied how small-scale societies manage climate variation. Here, we investigate how Yucatec Maya subsistence farmers respond to climate stress, and the ways in which market integration may enhance or disturb response stategies. Using information on harvest returns, climate perceptions, household economics and helping networks, modelling results show that as farmers rely more on market inputs (e.g. seed, tractors, fertilizer) for a successful yield, the reasons given for a bad harvest shift from climate variables to access to quality inputs. We also find that social and economic diversification is key to mediating a household's experience of climate and market shocks. The Maya are astute stewards of climate knowledge, and have effective social and economic means to mitigate potential fluctuations in food availability. In the transition from a subsistence to a market integrated economy, these traditional strategies become strained. Reliance on market inputs forges a more rigid food production system that conflicts with the diversity and flexibility on which traditional strategies depend to manage climate variation. Moving forward, the best policies would be those that facilitate maintaining an equal footing in both a subsistence maize economy, while incorporating new market opportunities. This article is part of the theme issue 'Climate change adaptation needs a science of culture'.

Associations between the concurrent use of clinical decision support and computerized provider order entry and the rates of appropriate prescribing at discharge.

INTRODUCTION: Electronic health record systems used in conjunction with clinical decision support (CDS) or computerized provider order entry (CPOE) have shown potential in improving quality of care, yet less is known about the effects of combination use of CDS and CPOE on prescribing rates at discharge. OBJECTIVES: This study investigates the effectiveness of combination use of CDS and CPOE on appropriate drug prescribing rates at discharge for AMI or HF patients. METHODS: Combination use of CDS and CPOE is defined as hospitals self-reporting full implementation across all hospital units of CDS reminders, CDS guidelines, and CPOE. Appropriate prescribing rates of aspirin, ACEI/ARBs, or beta blockers are defined using quality measures from Hospital Compare. Multivariate linear regressions are used to test for differences in mean appropriate prescribing rates between hospitals reporting combination use of CDS and CPOE, compared to those reporting the singular use of one or the other, or the absence of both. Covariates include hospital size, region, and ownership status. RESULTS: Approximately 10% of the sample reported full implementation of both CDS and CPOE, while 7% and 17% reported full use of only CPOE or only CDS, respectively. Hospitals reporting full use of CDS only reported between 0.2% (95% CI 0.04 - 1.0) and 1.6% (95% CI 0.6 - 2.6) higher appropriate prescribing rates compared to hospitals reporting use of neither system. Rates of prescribing by hospitals reporting full use of both CPOE and CDS did not significantly differ from the control group. CONCLUSIONS: Although associations found between full implementation of CDS and appropriate prescribing rates suggest that clinical decision tools are sufficient compared to basic EHR systems in improving prescribing at discharge, the modest differences raise doubt about the clinical relevance of the findings. Future studies need to continue investigating the causal nature and clinical relevance of these associations.

FSH dystrophy and a subtelomeric 4q haplotype: a new assay and associations with disease.

BACKGROUND: Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant disease associated with contraction of arrays of tandem 3.3-kb units (D4Z4) on subtelomeric 4q. Disease-linked arrays usually have fewer than 11 repeat units. Equally short D4Z4 arrays at subtelomeric 10q are not linked to FSHD. The newly described 4qA161 haplotype, which is more prevalent in pathogenic 4q alleles, involves sequences in and near D4Z4. METHODS: We developed two new assays for 4qA161, which are based upon direct sequencing of PCR products or detecting restriction fragment length polymorphisms. They were used to analyse single nucleotide polymorphisms (SNPs) indicative of 4q161 alleles. RESULTS: All (35/35) FSHD patients had one or two 4qA161 alleles (60% or 40%, respectively). In contrast, 46% (21/46) of control individuals had no 4qA161 allele (p<10(-4)), and 26% had homozygous 4qB163 alleles. CONCLUSIONS: Our results from a heterogeneous population are consistent with the previously described association of the 4qA161 haplotype with FSHD, but a causal association with pathogenesis is uncertain. In addition, we found that haplotype analysis is complicated by the presence of minor 10q alleles. Nonetheless, our sequencing assay for the 4qA161allele can enhance molecular diagnosis of FSHD, including prenatal diagnosis, and is simpler to perform than the previously described assay.

Why teams don't work. Interview by Diane Coutu.

The belief that teams make us more creative and productive--and are the best way to get things done--is deeply entrenched. But Hackman, a professor of organizational psychology at Harvard and a leading expert on teams, is having none of it. Research, he says, consistently shows that teams underperform despite all their extra resources. In an interview with senior editor Diane Coutu, Hackman explains where teams go wrong. Shockingly, most of the time members don't agree on what the team is supposed to be doing or even on who is on the team. The belief that bigger is better also compounds problems; as a team grows, the effort needed to manage links between members increases almost exponentially. Leaders need to be ruthless about defining teams and keeping them small (fewer than 10 members), and some individuals (like team destroyers) should simply be forced off. The leader also must set a compelling direction for the team--but in so doing, may encounter intense resistance that puts him or her at great risk. Hackman explores other fallacies about teams--for instance, that teams whose members have been together a long time become stale. In fact, research reveals that new teams make 50% more mistakes than established teams. To avoid complacency, though, every team needs a deviant--someone who is willing to make waves and open up the group to more ideas. Unfortunately, such individuals often get thrown off the team, robbing it of its chance to be magical. Leaders can't make a team do well. However, by being disciplined about how a team is set up and managed, instituting the right support systems, and providing coaching in group processes, they can increase the likelihood that a team will be great.

Using brain-based measures to compose teams: how individual capabilities and team collaboration strategies jointly shape performance.

Advances in understanding neural processes open the possibility of using brain-based measures to compose collaborative work teams. Neuroimaging studies have shown that individual differences in patterns of brain activity can predict differences in performance of specific tasks. We extended this finding by examining performance not simply by a single brain, but by pairs of brains. We used measures derived from brain-based studies to compose 100 two-person teams in which members' roles were either congruent or incongruent with their individual abilities. The assessed abilities are rooted in the visual system, which comprises independent "spatial" and "object" subsystems. The team task required one member to navigate through a virtual maze (a spatial task) and the other to remember "tag" repetitions of complex "greebles" (an object-properties task). Teams in which members' role assignments were congruent with their abilities performed better than incongruent teams and teams in which both members scored high on only one of the abilities. In addition, verbal collaboration enabled members of incongruent teams to overcome their compositional disadvantage but did not enhance the performance of congruent teams-and actually impaired performance in teams in which both members were adept in only one of the two necessary abilities. The findings show that knowledge about brain systems can not only be used to compose teams, but also provides insights into how teams can best perform.

Efaproxiral red blood cell concentration predicts efficacy in patients with brain metastases.

Efaproxiral (Efaproxyn, RSR13), a synthetic allosteric modifier of haemoglobin (Hb), decreases Hb-oxygen (O(2)) binding affinity and enhances oxygenation of hypoxic tumours during radiation therapy. This analysis evaluated the Phase 3, Radiation Enhancing Allosteric Compound for Hypoxic Brain Metastases; RT-009 (REACH) study efficacy results in relation to efaproxiral exposure (efaproxiral red blood cell concentration (E-RBC) and number of doses). Recursive partitioning analysis Class I or II patients with brain metastases from solid tumours received standard whole-brain radiation therapy (3 Gy/fraction x 10 days), plus supplemental O(2) (4 l/min), either with efaproxiral (75 or 100 mg/kg daily) or without (control). Efaproxiral red blood cell concentrations were linearly extrapolated to all efaproxiral doses received. Three patient populations were analysed: (1) all eligible, (2) non-small-cell lung cancer (NSCLC) as primary cancer, and (3) breast cancer primary. Efficacy endpoints were survival and response rate. Brain metastases patients achieving sufficient E-RBC (> or =483 microg/ml) and receiving at least seven of 10 efaproxiral doses were most likely to experience survival and response benefits. Patients with breast cancer primary tumours generally achieved the target efaproxiral exposure and therefore gained greater benefit from efaproxiral treatment than NSCLC patients. This analysis defined the efaproxiral concentration-dependence in survival and response rate improvement, and provided a clearer understanding of efaproxiral dosing requirements.

The role of titin in muscular disorders.

Titin, the biggest single (poly) peptide found in humans, and throughout nature so far, was long considered as a good candidate for inherited muscle diseases. However, disease-causing defects were not known until recently, when this central sarcomeric protein was associated with human skeletal tibial muscular dystrophy (TMD/LGMD2J), dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM). Several mutations in different parts of titin have now been identified and more are expected. Spontaneous mouse and zebrafish mutants have also been reported. Experimental knock-outs are not viable, even in cases where just a c-terminal part of the gene was silenced, telling something of the basic importance of titin for life. In this article we review the current known structure and functions of this elementary molecule with some emphasis on the only defects so far known to cause human skeletal muscle disease, mutations in the c-terminal M-line part of titin.

Human immunodeficiency virus-seronegative adults with extrapulmonary tuberculosis have abnormal innate immune responses.

Extrapulmonary tuberculosis is presumably a marker of underlying immunodeficiency, but cytokine response pathways in these patients have not been well studied. Cytokine responses of peripheral blood mononuclear cells from human immunodeficiency virus-seronegative adults with prior culture-confirmed extrapulmonary tuberculosis were compared with those of persons with latent Mycobacterium tuberculosis infection. Mitogen-stimulated interferon (IFN)-gamma production, interleukin (IL)-12 production, and IFN-gamma receptor- and IL-12 receptor-mediated cytokine production did not differ between case patients and control patients. However, median resting IL-8 production was significantly lower in case patients than control patients (8051 vs. 19,290 pg/mL; P=.009). In addition, the median tumor necrosis factor (TNF)-alpha response was lower in case patients than control patients after stimulation with lipopolysaccharide (833 vs. 1149 pg/mL; P=.06) and lipopolysaccharide plus IFN-gamma (3301 vs. 4411 pg/mL; P=.04). These abnormalities in resting IL-8 and lipopolysaccharide-induced TNF-alpha production were not associated with IFN-gamma or IL-12 abnormalities and were detected up to several years after cure of disease, suggesting an abnormality in innate immunity.

A randomized, controlled trial of interventions to improve adherence to isoniazid therapy to prevent tuberculosis in injection drug users.

PURPOSE: To determine the effect of several interventions on adherence to tuberculosis preventive therapy. METHODS: We conducted a randomized trial with a factorial design comparing strategies for improving adherence to isoniazid preventive therapy in 300 injection drug users with reactive tuberculin tests and no evidence of active tuberculosis. Patients were assigned to receive directly observed isoniazid preventive therapy twice weekly (Supervised group, n = 99), daily self-administered isoniazid with peer counseling and education (Peer group, n = 101), or routine care (Routine group, n = 100). Patients within each arm were also randomly assigned to receive an immediate or deferred monthly $10 stipend for maintaining adherence. The endpoints of the trial were completing 6 months of treatment, pill-taking as measured by self-report or observation, isoniazid metabolites present in urine, and bottle opening as determined by electronic monitors in a subset of patients. RESULTS: Completion of therapy was 80% for patients in the Supervised group, 78% in the Peer group, and 79% in the Routine group (P = 0.70). Completion was 83% (125 of 150) among patients receiving immediate incentives versus 75% (112 of 150) among patients with deferred incentives (P = 0.09). The proportion of patients who were observed or reported taking at least 80% of their doses was 82% for the Supervised arm of the study, compared with 71% for the Peer arm and 90% for the Routine arm. The proportion of patients who took 100% of doses was 77% for the Supervised arm (by observation), 6% for the Peer arm (by report), and 10% for the Routine arm (by report; P <0.001). Direct observation showed the median proportion of doses taken by the Supervised group was 100%, while electronic monitoring in a subset of patients showed the Peer group (n = 27) took 57% of prescribed doses and the Routine group (n = 32) took 49% (P <0.001). Patients in the Routine arm overreported adherence by twofold when data from electronic monitoring were used as a gold standard. There were no significant differences in electronically monitored adherence by type of incentive. CONCLUSION: Adherence to isoniazid preventive therapy by injection drug users is best with supervised care. Peer counseling improves adherence over routine care, as measured by electronic monitoring of pill caps, and patients receiving peer counseling more accurately reported their adherence. More widespread use of supervised care could contribute to reductions in tuberculosis rates among drug users and possibly other high-risk groups.

Pentobarbital-activated Cl(-) channels in cultured adult and embryonic human DRG neurons.

Developmental differences in pentobarbital-activated Cl(-) currents were studied in adult and embryonic human dorsal root ganglia (DRG) neurons using whole-cell patch-clamp recordings. Pentobarbital-induced Cl(-) conductance was significantly greater in adult DRGs (28.4 pS) than in embryonic cells (19.1 pS). Fluctuation analysis of the spectral density plots of Cl(-) channel activation by pentobarbital showed age differences in the length and number of open time constants (adult cells, tau(1), tau(2), tau(3) were 224, 8. 4, 1.5 ms, respectively; embryonic cells tau(1) and tau (2) were 165 and 26.3 ms, respectively). The different kinetic properties of human adult and embryonic DRG Cl(-) channels opened by pentobarbital may reflect the presence of different subunits in the two populations of neurons.

GABA-Induced Cl- current in cultured embryonic human dorsal root ganglion neurons.

gamma-Aminobutyric acid (GABA)-activated channels in embryonic (5-8 wk old) human dorsal root ganglion (DRG) neurons in dissociated culture were characterized by whole cell and single-channel techniques. All DRG neurons when held at negative holding membrane potentials displayed inward current to micromolar concentrations of GABA applied by pressure pulses from closely positioned micropipettes. The current was directly proportional to the concentration of GABA (EC50, 111 microM; Hill coefficient, 1.7). DRG neurons also responded to micromolar concentrations of pentobarbital and alphaxalone but not to cis-4-aminocrotonic acid (CACA), glycine, or taurine. Baclofen (100 microM) affected neither the holding currents nor K+ conductance (when patch pipettes were filled with 130 mM KCl) caused by depolarizing pulses. Whole cell GABA-currents were blocked by bicuculline, picrotoxin, and t-butylbicyclophosphorothionate (TBPS; all at 100 microM). The reversal potential of whole cell GABA-currents was close to the theoretical Cl- equilibrium potential, shifting with changes in intracellular Cl- concentration in a manner expected for Cl--selective channels. The whole cell I-V curve for GABA-induced currents demonstrated slight outward rectification with nearly symmetrical outside and inside Cl- concentrations. Spectral analysis of GABA-induced membrane current fluctuations showed that the kinetic components were best fitted by a triple Lorentzian function. The apparent elementary conductance for GABA-activated Cl- channels determined from the power spectra was 22.6 pS. Single-channel recordings from cell-attached patches with pipettes containing 10 microM GABA indicated that GABA-activated channels have a main and a subconductance level with values of 30 and 19 pS, respectively. Mean open and closed times of the channel were characterized by two or three exponential decay functions, suggesting two or three open channel states and two closed states. Single channels showed a lack of rectification. The actions of GABA on cultured human embryonic DRG neurons are mediated through the activation of GABAA receptors with properties corresponding to those found in the CNS of human and other mammalian species but differing from those of cultured human adult DRG neurons.

Alphaxalone activates a Cl- conductance independent of GABAA receptors in cultured embryonic human dorsal root ganglion neurons.

Whole cell and cell-attached patch-clamp techniques characterized the neurosteroid anesthetic alphaxalone's (5alpha-pregnane-3alpha-ol-11,20-dione) effects on GABAA receptors and on Cl- currents in cultured embryonic (5- to 8-wk old) human dorsal root ganglion neurons. Alphaxalone applied by pressure pulses from closely positioned micropipettes failed to potentiate the inward Cl- currents produced by application of GABA. In the absence of GABA, alphaxalone (0.1-5.0 microM) directly evoked inward currents in all dorsal root ganglion neurons voltage-clamped at negative membrane potentials. The amplitude of the current was directly proportional to the concentration of alphaxalone (Hill coefficient 1.3 +/- 0.15). The alphaxalone-induced whole cell current was carried largely by Cl- ions. Its reversal potential was close to the theoretical Cl- equilibrium potential, changing with a shift in the external Cl- concentration as predicted by the Nernst equation for Cl- ions. And because the alphaxalone-current was not suppressed by the competitive GABAA receptor antagonist bicuculline or by the channel blockers picrotoxin and t-butylbicyclophosphorothionate (TBPS; all at 100 microM), it did not appear to result from activation of GABAA receptors. In contrast to GABA-currents in the same neurons, the whole cell current-voltage curves produced in the presence of alphaxalone demonstrated strong inward rectification with nearly symmetrical bath and pipette Cl- concentrations. Fluctuation analysis of the membrane current variance produced by 1.0 microM alphaxalone showed that the power density spectra were best fitted to double Lorentzian functions. The elementary conductance for alphaxalone-activated Cl- channels determined by the relationship between mean amplitude of whole cell current and variance was 30 pS. Single-channel currents in cell-attached patches when the pipette solution contained 10 microM alphaxalone revealed a single conductance state with a chord conductance of approximately 29 pS. No subconductance states were seen. The current-voltage determinations for the single-channels activated by alphaxalone demonstrated a linear relationship. Mean open and shut times of single alphaxalone-activated channels were described by two exponential decay functions. Taken together, the results indicate that in embryonic human DRG neurons, micromolar concentrations of alphaxalone directly activate Cl- channels whose electrophysiological and pharmacological properties are distinct from those of Cl- channels associated with GABAA receptors.

Mechanisms involved in the metabotropic glutamate receptor-enhancement of NMDA-mediated motoneurone responses in frog spinal cord.

1. The metabotropic glutamate receptor (mGluR) agonist trans-(+/-)-1-amino-1,3-cyclopentanedicarboxylic acid (trans-ACPD) (10-100 microM) depolarized isolated frog spinal cord motoneurones, a process sensitive to kynurenate (1.0 mM) and tetrodotoxin (TTX) (0.783 microM). 2. In the presence of NMDA open channel blockers [Mg2+; (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK801); 3,5-dimethyl-1-adamantanamine hydrochloride (memantine)] and TTX, trans-ACPD significantly potentiated NMDA-induced motoneurone depolarizations, but not alpha-amino-3-hydroxy-5-methylisoxazole-4-proprionate (AMPA)- or kainate-induced depolarizations. 3. NMDA potentiation was blocked by (RS)-alpha-methyl-4-carboxyphenylglycine (MCPG) (240 microM), but not by alpha-methyl-(2S,3S,4S)-alpha-(carboxycyclopropyl)-glycine (MCCG) (290 microM) or by alpha-methyl-(S)-2-amino-4-phosphonobutyrate (L-MAP4) (250 microM), and was mimicked by 3,5-dihydroxyphenylglycine (DHPG) (30 microM), but not by L(+)-2-amino-4-phosphonobutyrate (L-AP4) (100 microM). Therefore, trans-ACPD's facilitatory effects appear to involve group I mGluRs. 4. Potentiation was prevented by the G-protein decoupling agent pertussis toxin (3-6 ng ml(-1), 36 h preincubation). The protein kinase C inhibitors staurosporine (2.0 microM) and N-(2-aminoethyl)-5-isoquinolinesulphonamide HCI (H9) (77 microM) did not significantly reduce enhanced NMDA responses. Protein kinase C activation with phorbol-12-myristate 13-acetate (5.0 microM) had no effect. 5. Intracellular Ca2+ depletion with thapsigargin (0.1 microM) (which inhibits Ca2+/ATPase), 1,2-bis(O-aminophenoxy)ethane-N,N,N',N'-tetracetic acid acetyl methyl ester (BAPTA-AM) (50 microM) (which buffers elevations of [Ca2+]i), and bathing spinal cords in nominally Ca2+-free medium all reduced trans-ACPD's effects. 6. The calmodulin antagonists N-(6-aminohexyl)-5-chloro-1-naphthalenesulphonamide (W7) (100 microM) and chlorpromazine (100 microM) diminished the potentiation. 7. In summary, group I mGluRs selectively facilitate NMDA-depolarization of frog motoneurones via a G-protein, a rise in [Ca2+]i from the presumed generation of phosphoinositides, binding of Ca2+ to calmodulin, and lessening of the Mg2+-produced channel block of the NMDA receptor.

Role of metabotropic glutamate receptors in the depression of GABA-mediated depolarization of frog primary afferent terminals.

Sucrose gap recordings from the dorsal roots of isolated, hemisected frog spinal cords were used to determine the effects of metabotropic L-glutamate receptor activation on primary afferent terminals by (+/-)-1-amino-trans-1,3-cyclopentane-dicarboxylic acid (t-ACPD). Dorsal root potentials evoked by ventral root volleys were significantly reduced by t-ACPD (30 microM), as were GABA- and muscimol-induced afferent terminal depolarizations. The effects of t-ACPD on GABA-depolarizations depended upon activation of group I metabotropic glutamate receptors, i.e. the effects were blocked by the group I/II antagonist (RS)-alpha-methyl-4-carboxyphenylglycine, but not by the group II antagonist alpha-methyl-(2S,3S,4S)-alpha-(carboxycyclopropyl)-glycine or the group III antagonist alpha-methyl-(S)-2-amino-4-phosphonobutyrate and were mimicked by the group I agonist 3,5-dihydroxyphenylglycine but were not mimicked by the group III agonist (S)-2-amino-4-phosphonobutyrate. Increasing the intracellular concentration of 3'-5'-cyclic adenosine monophosphate with 8-bromo-cAMP, forskolin, and 3-isobutyl-1-methylxanthine significantly reduced GABA depolarizations, but the protein kinase inhibitors Rp-adenosine 3,5-cyclic monophosphothioate triethylamine and N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide did not alter t-ACPD's depression of GABA depolarizations. The actions of t-ACPD on GABA depolarizations were neither mimicked nor blocked by phorbol-12-myristate 13-acetate, thapsigargin, staurosporine, or arachidonic acid, presumptive indications that the effects of t-ACPD did not involve phosphoinositide hydrolysis, the release of Ca2+ from intracellular stores, or the formation of arachidonate. t-ACPD's effects on GABA depolarizations were blocked by 20 mM Mg2+, the broad spectrum L-glutamate antagonist kynurenate, and the selective N-methyl-D-aspartate antagonist D(-)-2-amino-5-phosphonovaleric acid, but not by the non-N-methyl-D-aspartate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione. Low concentrations of N-methyl-D-aspartate (10 microM) mimicked the effect of t-ACPD on GABA responses. These results suggest that t-ACPD's depression of GABA depolarizations involves an indirect, three-stage mechanism that includes activation of Group I metabotropic glutamate receptors on interneurons and/or on afferent terminals, the release of L-glutamate from the latter structures, and the activation of N-methyl-D-aspartate receptors on primary afferent terminals. The depression of GABA depolarizations caused by the release of L-glutamate from afferent terminal and/or interneurons leads to a block of presynaptic inhibition (produced in the frog spinal cord by GABA) resulting in a positive feed-forward amplification of reflex transmission.

Pharmacologically novel GABA receptor in human dorsal root ganglion neurons.

1. Whole cell voltage-clamp studies of gamma-aminobutyric acid (GABA) receptors were performed on large (> 80 microns) cultured human dorsal root ganglion (DRG) neurons. 2. GABA and pentobarbital sodium when applied in micromolar concentrations evoked inward Cl- currents in DRG neurons voltage clamped at negative membrane potentials. 3. Diazepam (10 microM) and pentobarbital (10 microM) upmodulated the GABA current by approximately 149 and 168%, respectively. 4. The GABA currents in human DRG cells were unaffected by the classical GABA antagonists picrotoxin and bicuclline (100 microM). In contrast, the GABA responses evoked in adult rat DRG cells cultured in an identical manner were inhibited by both antagonists. The glycine receptor antagonist strychnine (100 microM) did not alter GABA currents in human DRG cells. 5. Human DRG cells did not respond to glycine (10-100 microM) or taurine (10-100 microM). The GABAB agonist baclofen had no effect on the holding current when patch pipettes were filled with 130 mM KCl. The GABAB antagonists saclofen applied either alone or with GABA was without effect. 6. The differences between the GABA receptors described here and GABA receptors in other species may reflect the presence of receptor subunits unique to human DRG cells.

Voltage-gated calcium currents in whole-cell patch-clamped bullfrog dorsal root ganglion cells: effects of cell size and intracellular solutions.

Acutely dissociated bullfrog dorsal root ganglion (DRG) cells could be divided into two classes by measurement of cell capacitance. A bimodal distribution of cell capacitance was found and a value of 75 pF was used to divide frog DRG cells into 'small' and 'large' types. Two distinct voltage-activated Ca2+ currents were evoked in both classes of cells: a rapidly inactivating, low-voltage-activated current and a slowly-inactivating, high-voltage-activated current. When the recording pipette contained CsCl, greater peak inward current values and densities were seen in large cells compared to small cells. No significant differences were observed in the distribution of low-and high-voltage-activated currents in small and large cells. Replacement of pipette solutions containing CsCl with solutions containing equimolar concentrations of Cs glutamate, L-arginine Cl, or N-methyl-D-glucamine significantly increased both the reversal potential and the maximum amplitude of the Ca2+ currents in both small and large DRG cells. These increases indicate that internal substitutions with organic ions suppresses outward currents more effectively than does CsCl. In contrast to findings with CsCl, when organic ions were used in the pipette solution a significantly higher proportion of low-threshold Ca2+ channels was observed in small cells compared to large cells. These observations indicate that when organic solutions were used internally, significant differences in the proportion of low-threshold to high-threshold Ca2+ channels were observed in small and large cells. The composition of the internal solution is a critical variable when determining the type and amount of inward Ca2+ current in different types of neurons.

Modulation of frog spinal cord interneuronal activity by activation of 5-HT3 receptors.

Motoneuron membrane potentials were recorded from the ventral roots of isolated, hemisected frog spinal cords using sucrose gap techniques. The effects of the selective 5-HT3 agonist 2-methyl-serotonin (2-Me-5HT) on the changes in motoneuron membrane potential produced by dorsal root stimulation and by superfusion of excitatory amino acid agonists were evaluated. Application of 2-Me-5HT (100 microM) did not alter motoneuron membrane potential, but did substantially reduce (approximately 20%) the polysynaptic ventral root potentials evoked by dorsal root stimulation. 2-Me-5HT did not change motoneuron depolarizations generated by addition to the Ringer's solution of the excitatory amino acid agonists AMPA (10-30 microM), kainate (30 microM), or t-ACPD (100 microM), but NMDA-induced motoneuron depolarizations (100 microM) were significantly and reversibly reduced (approximately 20%) by exposure to 2-Me-5HT (100 microM). 2-Me-5HT-evoked decreases of NMDA depolarizations were blocked by the 5-HT3 antagonists ICS 205 930 (50-100 microM) and D-tubocurarine (3-10 microM), but not by MDL 72222 (20-100 microM), the 5-HT2 receptor antagonist ketanserin (10 microM), or the 5-HT1A/5-HT2A antagonist spiperone (10 microM). Two lines of evidence support the hypothesis that the effects of 2-Me-5HT are generated by an indirect mechanism involving interneurons: (1) TTX (0.781 microM) eliminated the effect of 2-Me-5HT on NMDA-induced motoneuron depolarizations, and (2) 2-Me-5HT reduced spontaneous ventral root potentials that result from interneuronal discharges. We attempted to establish the identity of a putative transmitter released by interneurons responsible for the effects on NMDA-depolarizations produced by 2-Me-5HT, but the AMPA receptor antagonist, CNQX (10 microM), the GABAA receptor antagonist, bicuculline (50 microM), the GABAB receptor antagonist, saclofen (100 microM), the opioid antagonist, naloxone (100 microM), and the adenosine antagonists, CPT (20-100 microM) and CSC (10-100 microM) did not alter 2-Me-5HT-induced reductions of NMDA-depolarizations. In sum, the site of interaction between 2-Me-5HT and NMDA appears to be at interneuronal locus, but the mechanism remains unclear.

Changes in motoneuron membrane potential and reflex activity induced by sudden cooling of isolated spinal cords: differences among cold-sensitive, cold-resistant and freeze-tolerant amphibian species.

The effects of sudden cooling of the spinal cord were studied in three species of amphibians--a cold-sensitive tropical toad (Bufo marinus), a cold-resistant, aquatic, hibernating frog (Rana pipiens, northern leopard frog) and a freeze-tolerant frog (Rana sylvatica, wood frog). Ventral root (motoneuron) potentials were recorded from isolated, hemisected spinal cords of each species mounted in a sucrose-gap recording apparatus and superfused with HCO3(-)-buffered Ringer's solution at room temperature (21 degrees C). In the toad, sudden cooling to 6-8 degrees C produced large, sustained motoneuron depolarizations that returned slowly to baseline levels and were accompanied by extensive paroxysmal activity. Larger, but shorter-lasting, motoneuron depolarizations associated with only a limited amount of paroxysmal activity were generated by rapid cooling of the leopard frog spinal cord. Small, brief motoneuron depolarizations followed by a hyperpolarization, or hyperpolarizations not preceded by depolarizations, were seen in cooled wood frog spinal cords. The wood frog displayed a large amount of spontaneous motoneuron activity, but little paroxysmal activity in response to sudden cooling. Following prolonged cooling, rewarming the spinal cords of all three species resulted in motoneuron hyperpolarizations that slowly decayed towards the baseline value. The amplitude of the rewarming-induced response was larger and longer in toad motoneurons than in leopard frog and wood frog motoneurons. At room temperature, a single supramaximal dorsal root stimulus evoked a depolarizing ventral root potential in toad and leopard frog motoneurons that was decreased in amplitude and prolonged when the spinal cords were cooled to 8 degrees C or below. In contrast, at room temperature, the ventral root reflex in the wood frog was followed by a distinct hyperpolarization. Cooling the wood frog spinal cord only slightly reduced the amplitude of the ventral root potential. In contrast, the evoked hyperpolarization was blocked by sudden cooling and also by the addition of dihydro-ouabain to the Ringer's solution. The motoneuron hyperpolarizations induced by sudden cooling in the wood frog were converted to depolarizations when Cl- in the superfusate was replaced with isethionate. The depolarizations elicited by sudden cooling were reduced by the addition of kynurenate in all three species. A dose-response curve generated by short applications of L-glutamate demonstrated that wood frog motoneurons were less sensitive than leopard frog motoneurons to L-glutamate. In summary, three species of amphibians, differing in their adaptations to the temperature of their environments, vary in their responses to sudden reductions in temperature. The relationship of these responses to their environmental adaptations remains to be determined.

Properties of ibogaine and its principal metabolite (12-hydroxyibogamine) at the MK-801 binding site of the NMDA receptor complex.

The putative anti-addiction alkaloid ibogaine and its principal metabolite 12-hydroxyibogamine appear to act at the (+)-5 methyl-10,11,dihydro-5H- dibenzo[a,d]cycloheten-5-10-imine maleate (MK-801) binding site in the N-methyl-D-aspartate (NMDA)-receptor cation channel. This conclusion is based on findings that both compounds competitively displaced specific [3H]MK-801 binding to membranes from postmortem human caudate and cerebellum and from frog spinal cord. Ibogaine was 4-6-fold more potent than its metabolite and both compounds were less potent (50-1000-fold) than MK-801 binding to the NMDA receptor. In addition, ibogaine (100 microM) and 12-hydroxyibogamine (1 mM) blocked (85-90% of control) the ability of NMDA (100 microM, 5 s) to depolarize frog motoneurons in the isolated frog spinal cord. The prevention of NMDA-depolarizations in frog motoneurons showed use-dependency and was very similar to the block produced by MK-801. In view of the abilities of MK-801 to affect the responses to addictive substances in pre-clinical investigations, our results are compatible with the idea that the ability of ibogaine and 12-hydroxyibogamine to interrupt drug-seeking behavior may, in part, result from their actions at the MK-801 binding site.

Activation of alpha-adrenoceptors indirectly facilitates sodium pumping in frog motoneurons.

The effects of clonidine on Na+ pumping in motoneurons of the isolated frog spinal cord was investigated using sucrose gap recordings from ventral roots. Na+ pump activity, induced in motoneurons either by tetanizing the dorsal root or by rapidly exposing the cord to normal medium following 30 min in K(+)-free Ringer's solution (K(+)-activated hyperpolarizations), was increased by application of clonidine (100 microM). These actions of clonidine were blocked by the preferential alpha 2-adrenergic antagonist yohimbine, but not by alpha 1-adrenergic antagonist prazosin or the beta-blocker propranolol. Clonidine's effects on Na+ pumping appeared to be indirect (presumably via interneurons) because its effects on K(+)-activated hyperpolarizations were reduced by tetrodotoxin (TTX) or high concentrations of Mg2+. This indirect mechanism involved activation of non-NMDA excitatory amino acid receptors. Thus, in the presence of clonidine, CNQX, but not APH, limited the ability of clonidine to enhance K(+)-activated hyperpolarizations. The AMPA receptor may play a role in the process, K(+)-activated hyperpolarizations were augmented by the presence of AMPA; NMDA had no effect. The present results are consistent with the idea that activation of alpha 2-adrenoceptors produces the following: the release of excitatory amino acids from interneurons; the activation of non-NMDA receptors on motoneurons; increased Na+ influx and loading and increased Na+ pump activity.