Hemorheological and metabolic consequences of renal ischemia-reperfusion and their modulation by N,N-dimethyl-tryptamine on a rat model

Renal ischemia-reperfusion (I/R) involves interruption and restoration of blood flow and oxygenation, producing morphological and functional injury that depends on the duration of ischaemia and circumstances of reperfusion. It is relevant to many clinical contexts, notably transplantation where warm and cold ischaemia occur and reperfusion contributes to acute kidney injury, delayed graft function and chronic graft dysfunction. Previous research implicates metabolic disturbance, reactive oxygen species and inflammation in injury evolution; however, the micro-rheological consequences for red blood cells — specifically deformability and aggregation — have been incompletely characterised in renal I/R. Peto and colleagues set out to determine whether a 60-minute bilateral renal ischaemia followed by unilateral reperfusion alters metabolic and micro-rheological parameters in rats, and whether N,N-dimethyl-tryptamine (DMT), a sigma-1 receptor agonist with reported immunomodulatory and anti-inflammatory activity, can attenuate those changes. The study tested the hypothesis that DMT administration before ischaemia and before reperfusion would reduce I/R-induced deterioration in metabolic markers (for example lactate and acid-base balance) and in erythrocyte deformability and aggregation.

This experimental study used 26 healthy male Crl:WI rats (mean bodyweight 343.3 ± 29.5 g) randomly assigned to three groups: control (C, n = 6), ischaemia-reperfusion (I/R, n = 10) and I/R with DMT treatment (I/R+DMT, n = 10). Only males were used to avoid sex-related variability. Anaesthesia was induced with intraperitoneal thiopental (60 mg/kg) and subcutaneous atropine-sulphate (0.06 mg/kg). The protocol was approved by the University of Debrecen Committee of Animal Welfare and followed relevant national and EU regulations. Surgically, a midline laparotomy exposed both kidneys and the left femoral artery was cannulated for serial blood sampling. In the I/R protocol both renal pedicles were rendered ischaemic for 60 minutes; after that period the right kidney was excised and the left kidney reperfused for 120 minutes. In the I/R+DMT group DMT was administered intramuscularly 15 minutes before ligation/clamping (given as 2.95 ml/bwkg of a 2.45 mg/ml solution) and the dose was repeated 60 minutes later, 15 minutes before reperfusion. Animals were euthanized after the final sampling and tissue biopsies were taken for later histology. Blood samples (0.3 ml) were taken from the cannulated femoral artery at baseline (Base), immediately after clip removal (I-60), and at 30, 60 and 120 minutes of reperfusion (R-30, R-60, R-120). Equivalent timing was used for controls. Haematological variables (RBC, Hct, Hgb, MCV, MCH, MCHC, WBC, platelets) were measured on a Sysmex K-4500 analyser. Acid–base parameters, glucose and electrolytes (including pH, pO2, pCO2, bicarbonate, base excess, lactate, Na+, K+, Ca2+) were measured using an EPOC portable analyser. Red blood cell deformability was assessed by LoRRca MaxSis Osmoscan rotational ektacytometry (EI-SS curves, EI at 3 Pa, EImax, SS1/2 and EImax/SS1/2). Aggregation was measured by Myrenne MA-1 aggregometer with indices at 0 and 3 s-1 shear rates at 5 and 10 seconds (M5s, M10s, M1 5s, M1 10s). Statistical analysis used means ± SD with inter-group comparisons by Student's t-test or Mann–Whitney tests and intra-group comparisons by one-way or repeated measures ANOVA with appropriate post-hoc tests (Dunn's, Bonferroni, Student–Newman–Keuls), depending on distribution. Significance was defined as p < 0.05.

Overall, renal I/R produced metabolic derangement and micro-rheological impairment, and DMT attenuated but did not abolish these changes. Lactate rose markedly and blood pH fell in the I/R group; impaired erythrocyte deformability and enhanced aggregation were apparent from about the 30th minute of reperfusion. The I/R+DMT group showed smaller changes in lactate, pH, electrolytes and red blood cell rheology compared to untreated I/R animals. Haematology: In controls a slight decrease in RBC, Hgb and Hct occurred over time (RBC p = 0.012, Hgb p = 0.002, Hct p = 0.029 vs baseline). During reperfusion both I/R and I/R+DMT groups showed moderate haemoconcentration by R-120, with significant increases in Hgb and Hct versus control and/or baseline at various points (for example I/R Hgb p = 0.028 vs Control at R-120; I/R+DMT RBC p < 0.001 vs base and Control). Total WBC transiently rose in I/R immediately after the ischaemic period (p = 0.032 vs base, p = 0.016 vs Control) and later fell by 120 minutes (p = 0.013 vs base); WBC in I/R+DMT remained similar to controls. Platelet counts increased in I/R during early reperfusion (30 and 60 minutes, p < 0.001 vs base); conversely platelets were mildly decreased in the I/R+DMT group at several time points versus control and I/R. Metabolic parameters: Blood pH fell more in I/R than in controls by the end of the experiment; in the I/R+DMT group pH showed a moderate increase versus baseline (p = 0.029). pO2 decreased in Control and I/R at R-120 but remained steady in I/R+DMT (R-120 p = 0.013 vs Control). pCO2 and bicarbonate fell in all groups by R-120, most markedly in I/R. Base excess became negative in I/R and I/R+DMT groups (I/R p < 0.001 vs base). Lactate rose markedly in I/R (p < 0.001 vs base) and to a lesser but significant extent in I/R+DMT (p = 0.045 vs base); the control rise was not significant. Glucose decreased significantly only in I/R+DMT (p = 0.045 vs base). Na+ was stable, while K+ increased in all groups and most in I/R; I/R+DMT K+ increases reached significance versus base and versus Control and I/R at some points. Ca2+ showed a mild but significant decrease in all groups. Red blood cell deformability: EI at 3 Pa remained stable in controls but decreased in I/R at the end of bilateral ischaemia and during reperfusion (I-60 p = 0.007 vs Control; R-30 p = 0.035; R-60 p = 0.002 and p = 0.028 vs base). The I/R+DMT group had higher EI than the I/R group at multiple time points (I-60 p = 0.039; R-30 p = 0.003; R-60 p < 0.001; R-120 p = 0.001). The composite EImax/SS1/2 ratio, an indicator favouring deformability when larger, was lowest in I/R at several time points (for example I-60 p = 0.008 vs Control and p < 0.001 vs I/R+DMT) indicating impaired deformability that was mitigated by DMT. Red blood cell aggregation: Aggregation indices showed complex, index-specific patterns. M5s rose initially in all groups; in controls the rise persisted transiently and then normalised, in I/R the values decreased after ischaemia, while in I/R+DMT indices increased further during reperfusion with significant differences versus control and versus I/R at R-30 and R-60 (for example R-30 p < 0.001 vs base, p = 0.011 vs Control, p < 0.001 vs I/R). M1 5s displayed time-dependent changes with the largest relative rise in I/R+DMT. M10s decreased in controls over time but increased in I/R (I-60 p = 0.006 vs Control; R-30 p < 0.001 vs Control); the I/R+DMT group showed a later rise peaking at R-30 (p = 0.012 vs Control, p = 0.041 vs I/R). Overall, aggregation measures were inconsistent across indices, but the investigators reported enhanced aggregation and impaired deformability from roughly the 30th minute of reperfusion, with DMT altering the dynamics and magnitude of those changes.

Peto and colleagues interpret their results as evidence that 60 minutes of bilateral renal ischaemia followed by unilateral reperfusion produces metabolic disturbance and worsened micro-rheological parameters, changes they link to oxidative stress, inflammation and metabolic derangement during I/R. The authors describe mechanisms by which free radicals, leukocyte activation and ATP depletion alter erythrocyte membrane and shape, leading to reduced deformability and altered aggregation, and they relate the observed rises in haematocrit and lactate and the pH fall to these processes. The study team notes that hemorheological data in renal I/R are scarce compared with other organs. They contrast their findings with a previous clinical report in living-donor transplantation that reported reduced viscosity and improved deformation after transplantation, suggesting that haemorreological responses may differ by context and that improving microcirculation can be protective. Against that background, the investigators evaluated DMT because of recent reports that dimethyltryptamines act as immune regulators via the sigma-1 receptor. They report that DMT, given before ischaemia and before reperfusion, effectively reduced the magnitude of I/R-induced changes in several metabolic and rheological parameters, although it did not fully prevent them. The authors therefore propose that the DMT–sigma-1 axis is a promising target for modulating inflammation and limiting I/R injury. Limitations acknowledged by the researchers include the restricted blood sample volumes, which prevented use of syllectometry that could have provided a higher-resolution view of the early aggregation dynamics; they note that aggregation in rats is rapid and that their light-transmission measures offer only snapshots at 5 and 10 seconds. The authors also imply that index-specific and time-dependent variability in aggregation complicates interpretation. Finally, they observe the absence of prior literature on DMT use in renal I/R and frame their findings as preliminary evidence of a modulatory effect rather than proof of clinical utility.

Metabolic and micro-rheological parameters deteriorated after 60 minutes of bilateral renal ischaemia and during subsequent unilateral reperfusion in this rat model. Intramuscular N,N-dimethyl-tryptamine administered before ischaemia and before reperfusion reduced but did not completely prevent changes in lactate, pH, electrolytes, red blood cell deformability and aggregation.