The Making of a Capital: Amman, 1918-1928

Par deux fois dans son histoire moderne, Amman s’est imposée aux dépens de centres administratifs plus anciens: al-Salt dans les premières années de l’émir Abdallah, et Jérusalem entre les guerres de 1948 et 1967. Cet article s’intéresse au premier cas, couvrant les années qui séparent le retrait ottoman en 1918 et la désignation formelle de Amman comme capitale de l’Emirat de Transjordanie en 1928. La formation d’un Etat colonial sert de toile de fond à l’étude des dimensions institutionnelles de la formation d’une capitale: instances municipales et gouvernementales, infrastructures et usages de l’espace destinés à renforcer l’autorité de l’Etat, etc. La logique sous-jacente à la création d’une nouvelle ville pour accompagner celle d’un nouvel Etat permet de comprendre le choix de l’émir Abdallah pour cette ville circassienne de préférence à la vieille capitale ottomane du district, al-Salt.Twice in its modem history, Amman has expanded at the expense of older administrative centres: from al-Salt in the early years of Emir Abdullah’s reign, and from Jerusalem in the years between the 1948 and 1967 wars. This paper focuses on the earlier case, spanning the years between the Ottoman withdrawal in 1918 and the formal designation of Amman as the capital of the Emirate of Transjordan in 1928. The institutional meaning of a capital city is studied against the backdrop of the formation of a colonial state: the establishment of municipal and government offices, infrastructure and the use of space to reinforce the state’s authority. The underlying logic of creating a new city to accompany the process of creating a new state extends our understanding of Emir Abdullah’s preference for the Circassian new town over al-Salt, the old Ottoman district capital

High-throughput electrophysiological assays for voltage gated ion channels using SyncroPatch 768PE

<div><p>Ion channels regulate a variety of physiological processes and represent an important class of drug target. Among the many methods of studying ion channel function, patch clamp electrophysiology is considered the gold standard by providing the ultimate precision and flexibility. However, its utility in ion channel drug discovery is impeded by low throughput. Additionally, characterization of endogenous ion channels in primary cells remains technical challenging. In recent years, many automated patch clamp (APC) platforms have been developed to overcome these challenges, albeit with varying throughput, data quality and success rate. In this study, we utilized SyncroPatch 768PE, one of the latest generation APC platforms which conducts parallel recording from two-384 modules with giga-seal data quality, to push these 2 boundaries. By optimizing various cell patching parameters and a two-step voltage protocol, we developed a high throughput APC assay for the voltage-gated sodium channel Nav1.7. By testing a group of Nav1.7 reference compounds’ IC₅₀, this assay was proved to be highly consistent with manual patch clamp (R > 0.9). In a pilot screening of 10,000 compounds, the success rate, defined by > 500 MΩ seal resistance and >500 pA peak current, was 79%. The assay was robust with daily throughput ~ 6,000 data points and Z’ factor 0.72. Using the same platform, we also successfully recorded endogenous voltage-gated potassium channel Kv1.3 in primary T cells. Together, our data suggest that SyncroPatch 768PE provides a powerful platform for ion channel research and drug discovery.</p></div

SyncroPatch cell patching success rate optimization.

<p>(A) Cell harvesting with accutase brought better cell catching rate than Trypsin, and 1 time cell washing with PBS significantly improved cell catching rate; (B) Increasing Cell number up to 2000 per well reached the best cell catching rate; (C) The best cell membrane breaking-in pressure for our tested Nav1.7 and Nav1.5 cell lines were at -250 mBar; (D) The distribution of R_SEAL from optimized Nav1.7 and Nav1.5 recordings; (E) Under optimized cell patching parameters, SyncroPatch CHO-Nav1.7 and CHL-Nav1.5 cell patching success rate by each criterion and all criteria. Note that all comparison experiments were done by fixing other parameters at the optimized condition and varying the experimental parameter only; (F) Under optimized APC parameters, voltage gated sodium current recording success rate from Nav1.1, Nav1.2, Nav1.3, Nav1.4, Nav1.5, Nav1.6, Nav1.7 and NG108-15 cell line was 66%, 51%, 54%, 82%, 75%, 59%, 79% and 35%, respectively; (G) A representative SyncroPatch recording of CHO-Nav1.7 (left half chip) and CHL-Nav1.5 (right half chip) in a 384 well chip. The R_SEAL in each well was indicated as less than 200 MΩ in gray, between 200 MΩ and 1 GΩ in blue, and bigger than 1 GΩ in green by using SyncroPatch PatchControl software; All data shown as mean ± SD, with data points in SyncroPatch n = 200~384.</p

Acceptance criteria for automated patch clamp.

<p>Acceptance criteria for automated patch clamp.</p

SyncroPatch Nav1.7 recording from optimized conditions with two-state voltage protocol.

<p>(A) Voltage protocol to elicit Nav1.7 currents from closed (-120 mV) and inactivated (-40 mV) states. (B) Representative single sweep Nav1.7 current under described voltage protocol. The closed and inactivated state elicited peak currents were auto selected between red and green cursors. (C) The same representative recording current time plot shows steady closed state (dark blue) and inactivated state (light blue) peak currents through the whole experiment, including 3 times external solution washing (W1, W2 and W3) and 10 mins after applying 0.2% DMSO external solution.</p

Comparison of SyncroPatch APC and manual patch clamp (MPC) by characterizing Nav1.7, Nav1.5 and Kv1.3.

<p>(A) Representative Nav1.7 current traces from APC. The currents were elicited by 20 ms test pulses (-80 to 40 mV in 5 mV increment) from a holding potential at -120 mV. (B) Overlay of Nav1.7 IV relationship curves from APC and MPC. The peaks of the IV from both systems were -10 mV. (C) Superimposition of steady-state activation and inactivation curves of Nav1.7 from APC and MPC. Inactivation currents were elicited by 20 ms test pulses at -10 mV, after 500 ms conditioning prepulses ranged from—120 to 0 mV with 5 mV increments. The smooth curves are Boltzmann fits with activation V½ and slope factors (k) from APC -27.3 ± 0.9 mV and 2.6 ± 0.2 mV/e-fold potential change; and from MPC -26.1 ± 1.6 mV and 2.4 ± 0.3 mV/e-fold potential change. For inactivation the V½ and k values are -69.4 ± 0.4 mV and 8.9 ± 0.3 mV/e-fold potential change from APC and -69.7 ± 0.7 mV and 8.3 ± 0.6 mV/e-fold potential change from MPC. (D) Peak current (elicited by -10 mV 20 ms) was plotted as a function of inter-stimulus interval (prepulse and holding Vm at -120mV) ranging from 1 ms to 1,000 ms, and fitted with one phase decay exponential equation to obtain the recovery time constant, τ = 1.85 ± 0.1 and 3.65 ± 0.2 ms from APC and MPC, respectively. (E) Representative Nav1.5 current traces from APC. (F) Superimposition of Nav1.5 steady-state activation and inactivation curves from APC and MPC. The smooth curves are Boltzmann fits with activation V½ and slope factors (k) are -50.3 ± 0.5 mV and 2.7 ± 1.0 mV/e-fold potential change from APC; and are -50.1 ± 0.4 mV and 3.5 ± 0.5 mV/e-fold potential change from MPC. For inactivation the V½ and k values are -72 ± 0.4 mV and 5.9 ± 0.4 mV/e-fold potential change from APC; and are -73.5 ± 0.7 mV and 5.8 ± 0.6 mV/e-fold potential change from MPC. (G) Representative Kv1.3 current traces from APC. (H) Superimposition of Kv1.3 steady-state activation and inactivation curves from APC and MPC. The smooth curves are Boltzmann fits with activation V½ and slope factors (k) are -27.9 ± 1.0 mV and 8.5 ± 0.4 mV/e-fold potential change from APC; and are -29.4 ± 1.6 mV and 7.9 ± 1.4 mV/e-fold potential change from MPC. For inactivation the V½ and k values are -44.5 ± 0.5 mV and 4.1 ± 0.4 mV/e-fold potential change from APC; and are -45.2 ± 0.5 mV and 4.2 ± 0.4 mV/e-fold potential change from MPC. Note that all normalized data were shown as mean ± SEM, with data points in APC n = 290 ~ 384 and MPC n = 4 ~ 10.</p