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Histological studies and magnetic resonance imaging were employed to analyze the kidney structure and function of the freshwater mussel, Nodularia douglasiae. The Nodularia kidney consists of proximal, intermediate and distal tubules. The epithelia of the renal tubules were composed of a single layer of cuboidal cells. The proximal and distal tubules run in opposite directions underneath the pericardial cavity. Molecular weight cut-off (MWCO) values for the kidney filtration were detected by MR tracer injections: gadolinium-diethylenetriaminepentaacetic acid (GdDTPA) at 0.55 kDa, an oligomer-based contrast agent (CH3-DTPA-Gd) at 2.2 kDa, as well as Gd-DTPA-polylysine at 10, 22, and 110 kDa. The T1w-MRI intensity and T1 relaxation rate (R1) of the pericardial cavity and renal tubules increased with tracers smaller than 10 kDa. The other tracers showed only minimal or no increase. Thus, we concluded that the MWCO of the kidney is 22 kDa, 50 times larger than that for the Mytilus living in seawater. Since the R1 values of the renal tubules were similar to those of the pericardial cavity, the kidney did not concentrate filtrated tracers. The slow decay of the magnetic resonance (MR) tracers from the renal tubules indicated a low filtration rate, suggesting that the counter-current system reabsorbs useful solutes without reabsorption of water. The higher MWCO may be beneficial to maintain the tubular oncotic pressure and allow excretion of excess water. In conclusion, a main renal function of the freshwater mussel is the excretion of water, opposite to that of the seawater mussel and vertebrates, which preserve water., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

In order to determine the molecular weight cut-off (MWCO) for the atrial wall filtration into kidneys of the Mytilus galloprovincialis , we employed five magnetic resonance (MR) tracers: manganese chloride (Mn 2+ ), gadolinium chloride (Gd 3+ ), manganese-ethylenediaminetetraacetic acid (MnEDTA), gadolinium-diethylenetriamine pentaacetic acid (GdDTPA) and oligomer-based contrast agent (CH3-DTPA-Gd). After injection of the MR tracers (1 or 2 mmol l -1 ×0.1 ml) into the visceral mass, T 1 -weighted MR imaging ( T 1w ) were measured at 20°C. The MR tracers were distributed uniformly in the visceral mass within 1 h after injection. The T 1 of the kidney ( R 1 ) were measured at 20°C. The MR tracers were distributed uniformly in the visceral mass within 1 h after injection. The T 1w -MRI intensity and R 1 of the kidney ( R 1K ) were increased by Mn 2+ and MnEDTA, with urine concentrations estimated at 210 and 65 µmol l -1 , respectively. The rest of the tracers showed only minimal or no increase. When the mussels were additionally incubated in seawater with 10 µmol l -1 MnCl 2 , R 1K was increased in the GdDTPA group, but not in the GdCl 3 group. Therefore, Gd 3+ might have inhibited renal accumulation of Mn 2+ and Gd 3+ Incubation in seawater with 10 µmol l -1 MnEDTA showed no increase in the R 1K , but additional incubation with 10 µmol l -1 MnCl 2 caused an increase in R 1K It is suggested that injected MnEDTA was filtrated as MnEDTA per se, and not likely separated into free Mn 2+ Thus, we concluded that the MWCO of the atrial wall of the M. galloprovincialis is around 0.5 kDa, which is almost 1/100 of that for vertebrate animals, and suggests a reduction in efforts to reabsorb metabolites and osmolytes from the urine., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)