PHYSI'OLOGIA PLANTARUM 91: 477^87. 1994 Copyriglu © Phy^udi^m Plamirmn t994 Primed in Dennuifk — aSi H^hsx fcierved Evidence that short-term regulation of nodule permeahility does not occur in the inner cortex Owen R. Van Cauwenberghe, Stephen Hunt, William Newcomb, Martin J. Canny and David B. Layzell Van Cauwenberghe, O. R., Htint, S., Newcomh, W,, Canny, M,J. and Layzell, D'. B, 1994, Evidence thai short-term regulation of nodule permeahility does not occur in the inner cortex, - Pbysiol, Plam. 91: 477-487, Regulation of nodule permeability m response to short-term changes in environmental and physiological conditions is thought to occor by occlusion of intercellular spaces in tbe nodule inner cortex. To test this bypothesis, the permeability of legume Bodules was altered hy adapting them to either 20 or 80% O; over a 2.5-h period. The nodules were tben rapidly frozen, cryo-planed and examined under ciyo-scanning electron mi- croscopy for difference.? in the nonoher, area or sbape factor of intercellular spaces. Comparisons were made between whole nodules and specific nodule zone.s (outer cortex, middle cortex, inner cortex and central zone) in eacb treatment. Gas analysis niiea,s,urements indicated tbat nodules equilibrated at 20% O, bad a 6.6-fold bigher penmeability than those equilibrated at 80% O,. However, no significant differences were observed between pO, treatments in tbe number of open intercellular spaces, tbe cross-sectional area of those spaces, or the proportion of tbe tissue area present as open space in wbole nodules or any nodule zone. Also, altbougb nodules in both treatments posse.ssed a boundary layer of tigbtly packed cells in tbe inner cortex, tbe total ai-ea of intercellular spaces between cells bordering tbis layer did not differ hetween treat- ments,. Togetber tbese observations dO' not support tbe cun'ently favored hypothesis that nodule permeability is regulated hy opening or occlusion of intercellulai- spaces in the nodule inner cortex. Highly significant differences (P=0.0006) were observed be- tween '0'; treatments in tbe shape factor of the open intercellulai' spaces in all nodule zones. Nodules equilibrated at 80% OT had significantly more isodiametric spaces while those equilibrated at 20% O:, had more long, narrow spaces. This observation suggests that the critical step in the regulation of nodule permeahility to OT may be localized in tbe central, infected zone and involve changes in the ratio of the surface area of tbe intercellular space to tbe volume of the infected cell. Key words - Cryo-planing. cryo-SEM, diffusion barrier, intercellulai- space, oxygen concentration, permeability, shape factor, soyhean root nodules, O, R, Vati Cauwenberghe, S, Himt, W, Newcomb and D, B, Layzell (correspotiditig author), Dept of Biology, Queen's Utilv,. Kittgston, ON, Canada, K7L3N6; M,J. Catiny, Dept of Biology, Caiieton Univ,, Ottawa, ON, Canada, KIS5B6. . (Witty et al. 1987) have shown that the OT concentration '"' *"* drops very rapidly from atmospheric levels in the nodule Experitnental evidence indicates that a variable barrier to 'Outer cortex to levels helow the sensitivity 'Of the elec- gas diffusion and a high rate of O2 consumption work trode within the central zone. Thi.s has led to the hypothe- together to maintain a low O2 'Concentration in the central sis that the site of rtodule O2 regulation is located in the zone of legume nodules (Hunt et al. 1987, Weisz and nodule inner cottex (Willy et al. 1984, 1987). This hy- Sinclair 1988). Studies in which O2 microelectrodes have pothesis is supported by tnathemadcal models of gas been inserted into nodules of soybeati (Tjepkema and diffusion (Sheehy et ai. 1987, Hunt et al. 1988, Denison Yocum 1974, Masepohl et al. 1993), pea and french bean 1992) and by the presettce of a cell layer in the inner Received 16 August, 1993; revised 10 February, 1994 32 Physiol, Ptanc, St. 1994 477