Does salinity make a difference—Kidney anatomy of Saimaa (Pusa saimensis) and Baltic ringed seals (Pusa hispida botnica)

Abstract

As habitat salinity markedly differs between the endangered, freshwater‐dwelling Saimaa ringed seal (Pusa saimensis Nordquist, 1899) and the brackish water‐inhabiting Baltic ringed seal (Pusa hispida botnica Gmelin, 1788), we investigated whether this difference has resulted in morphological changes to their kidneys. We studied kidney anatomy using dissection, silicone casts, and standard histological methods. The Saimaa ringed seal had statistically significantly fewer reniculi than the Baltic ringed seal, and the mean number of reniculi was significantly smaller in both studied species than that previously reported from Arctic ringed seals (Pusa hispida hispida Schreber, 1775). We described four of the most common branching patterns of the renal artery. Contrary to previous findings in other ringed seal studies, we observed veins resembling arcuate and interlobar veins in histological sections of both species.

1. INTRODUCTION

The multilobed kidneys of pinnipeds and cetaceans are reniculate, consisting of numerous reniculi, each corresponding to a lobe or a unilobar kidney. As the reniculate structure does not allow greater urine concentration abilities, reniculation is believed to have evolved in response to the large body size of these animals (Bester, 1975; Ortiz, 2001; Vardy & Bryden, 1981), enabling relatively more glomeruli (Harrison and Tomlinson, 1956; Vardy & Bryden, 1981) and shorter tubules in a large kidney (Maluf & Gassman, 1998). The quantification of reniculi is challenging in pinnipeds due to the lack of interrenicular connective tissue and variation in their number between species (Vardy & Bryden, 1981; Williams & Worthy, 2002) and, to some extent, between individuals of the same species (Dragert et al., 1975; Vardy & Bryden, 1981). Data from previous studies indicate that the reniculi number correlates with species size (Dragert et al., 1975; Munkacsi & Newstead, 1985; Slijper, 1958; Vardy & Bryden, 1981; Williams & Worthy, 2002), and Slijper (1958) and Williams and Worthy (2002) noted that species with higher‐salinity diets appear to have more reniculi. Torres‐Romero et al. (2016) further suggested that larger body size provides better osmoregulatory abilities, leading to a positive correlation between body size and habitat salinity. Of the ringed seal subspecies, Munkacsi and Newstead (1985) reported around 140 reniculi in the kidney of an Arctic ringed seal (Pusa hispida hispida Schreber, 1775).

Besides interspecific differences in the number of reniculi, differences have been reported in the circulatory anatomy of the kidneys between pinniped species (Dragert et al., 1975; Guzsal, 1959; Harrison & Tomlinson, 1956; Munkacsi & Newstead, 1985; Van der Spoel, 1963; Vardy & Bryden, 1981). In phocid seals, the venous circulation of a reniculus is mostly or entirely directed peripherally, resulting in a great venous plexus surrounding the kidneys, which drains to the vena cava caudalis via multiple large veins (Guzsal, 1959; Munkacsi & Newstead, 1985; Van der Spoel, 1963; Vardy & Bryden, 1981). Arcuate veins, which in terrestrial mammals collect the venous blood from interlobular veins and drain to interlobar veins, are reportedly absent in ringed seals (Guzsal, 1959; Munkacsi & Newstead, 1985) and harp seals (Pagophilus groenlandicus Erxleben, 1777) (Dragert et al., 1975). However, arcuate veins have been described in harbor seals (Phoca vitulina Linnaeus, 1758) (Van der Spoel, 1963), crabeater seals (Lobodon carcinophaga Hombron & Jacquinot, 1842), and Weddell seals (Leptonychotes weddellii Lesson, 1826) (Vardy & Bryden, 1981). In otariids, Cape fur seal's (Arctocephalus pusillus Schreber, 1775) kidneys possess both intrinsic and extrinsic venous pathways, which, differing from phocids, drain to a vena renalis (Bester, 1975).

Of freshwater seals, the anatomy of the kidneys has been previously studied in the Ladoga ringed seal (Pusa hispida ladogensis Nordquist, 1899) by Guzsal (1959). His findings corresponded with those reported from the Arctic ringed seal (Munkacsi & Newstead's, 1985), but Guzsal (1959) did not report the number of reniculi of the studied Ladoga ringed seal individual. The Saimaa ringed seal (Pusa saimensis, previously Pusa hispida saimensis Nordquist, 1899) is an endangered, freshwater seal species, which was until recently reckoned as a subspecies of the ringed seal. The population has been isolated from its geographically closest related populations, the Baltic ringed seals (Pusa hispida botnica Gmelin, 1788) and Ladoga ringed seals in Lake Saimaa, Finland, for at least since the end of the last glaciation period (Ukkonen et al., 2014; Valtonen et al., 2012). Genetically, the Saimaa ringed seal differs significantly from ringed seal subspecies (Löytynoja et al., 2023; Martinez‐Bakker et al., 2013) with an evolutionary history extending beyond the last glaciation (Löytynoja et al., 2023, 2025). In anatomical studies, minor morphological differences have been found between the Saimaa ringed seal and the Baltic ringed seal in skull morphology (Amano et al., 2002; Laakkonen & Jernvall, 2020; Nordqvist, 1899), the lower respiratory tract (Laakkonen & Jernvall, 2016), the temporomandibular joint (Laakkonen & Jernvall, 2020), and the tongue and teeth (Löytynoja et al., 2025). Taken together, these genetic and anatomical differences were recently used to propose a species‐level rank for the Saimaa ringed seal (Löytynoja et al., 2025). Consequently, the Society of Marine Mammology recognized the Saimaa ringed seal as a distinct species in 2025 (https://marinemammalscience.org/facts/p-h-saimensis/).

The Baltic Sea, a brackish waterbody, was colonized by the Baltic ringed seal over 10,200 years ago (Ukkonen et al., 2014). The population of more than 20,000 individuals (Ersalman et al., 2025) is believed to have retained a connection to the Arctic ringed seal population (Martinez‐Bakker et al., 2013; Palo et al., 2001; Ukkonen et al., 2014; Valtonen et al., 2012). Despite some anatomical differences (Amano et al., 2002), the subspecies status of the Baltic ringed seal has been questioned due to genetic similarity between Baltic and Arctic ringed seals (Berta & Churchill, 2012; Martinez‐Bakker et al., 2013; Palo et al., 2001). Nevertheless, more recent analyses suggest retention of distinct status for the Baltic population (Olsen et al., 2025).

The aim of our study was to compare the kidney anatomy of the Saimaa and Baltic ringed seals to that of their marine counterpart, the Arctic ringed seal, to determine whether the difference in salinity is reflected in their kidney anatomy. Our hypothesis was that the lower environmental salinity of both the Saimaa and Baltic ringed seal habitats compared to that of marine ringed seals has resulted in changes to kidney anatomy.

2. MATERIALS AND METHODS

All Saimaa ringed seals found dead are collected by the Finnish governmental institute Metsähallitus. The carcasses are stored at −20°C for pathological examinations carried out at the Finnish Food Authority in Oulu, Finland. After measurements and necropsy, various samples were collected for biological research purposes. Metsähallitus and the University of Helsinki share a research agreement that permits the use of Saimaa ringed seal material in research. Most Saimaa ringed seal material used in this study was collected during these examinations in 2016–2024. In addition, during 2018–2020, six Saimaa ringed seals were anatomically dissected at the Faculty of Veterinary Medicine in the University of Helsinki. These animals had died in gill nets and pathological examinations were therefore unnecessary. After examinations in Oulu and Helsinki, the lower canine tooth of each animal was sent to Natural Resources Institute Finland for age evaluation (Stewart et al., 1996).

Histological samples were also taken from a Saimaa ringed seal pup (specimen number 2755), which was found malnourished in July 2019 and taken to the Wildlife Hospital of Korkeasaari Zoo (Metsähallitus, 2020). The pup died of intestinal obstruction at the Wildlife Hospital in February 2020 (Metsähallitus, 2020). The carcass was taken to the Faculty of Veterinary Medicine for pathological examination and stored at +6°C. Histological samples were taken within a day after the death.

Six Baltic ringed seal carcasses were obtained from a hunter from the Gulf of Bothnia during 2018–2020. After death, carcasses were frozen and transported to Helsinki, where they were examined at the Faculty of Veterinary Medicine of the University of Helsinki. In addition, one or both kidneys of seven Baltic ringed seal individuals were received from the Natural Resources Institute Finland. These kidneys were surplus material, sent to the Natural Resources Institute Finland by hunters along with other samples. Consequently, the weight information of these seals was mostly estimates recorded in the field. Baltic ringed seals are legally hunted in Finland, and the studied individuals were hunted for reasons unrelated to this research. In total, one or both kidneys or kidney samples from 28 Saimaa ringed seals and 14 Baltic ringed seals were used in this study (Table 1).

TABLE 1.

Specimens used for examining the circulatory and urethral anatomy of Saimaa ringed seals (n = 19) and Baltic ringed seals (n = 14), identified by specimen numbers issued by Metsähallitus for the Saimaa ringed seals and by Natural Resources Institute Finland for the Baltic ringed seals. In total, 25 kidneys from Saimaa ringed seals and 20 from Baltic ringed seals were examined.

Species	Identification	Age group	Sex	Left/right	Ureter	Renal artery	Renal veins
Saimaa ringed seal	2664	Pup	M	Right		x
Saimaa ringed seal	2670	Subadult	F	Left			x
Saimaa ringed seal	2673	Pup a	M	Left		x	x
	2673			Right		x	x
Saimaa ringed seal	2697	Pup	M	Left			x
	2697			Right
Saimaa ringed seal	2698	Subadult	M	Left	x		x
Saimaa ringed seal	2702	Subadult	M	Left	x	x
	2702			Right		x
Saimaa ringed seal	2703	Pup	M	Right	x
Saimaa ringed seal	2705	Subadult a	F	Left		x	x
	2705			Right		x
Saimaa ringed seal	2728	Pup	F	Left	x	x
Saimaa ringed seal	2734	Pup a	F	Right	x	x
Saimaa ringed seal	2742	Pup	F	Right	x	x
Saimaa ringed seal	2743	Pup a	M	Left	x	x
Saimaa ringed seal	2744	Pup a	F	Left	x	x
	2744			Right	x	x
Saimaa ringed seal	2745	Pup a	F	Left	x
	2745			Right		x
Saimaa ringed seal	2747	Pup a	F	Right	x	x
Saimaa ringed seal	2754	Subadult a	F	Left	x	x
Saimaa ringed seal	2777	Adult	M	Left		x
Saimaa ringed seal	2778	Pup a	F	Right		x
Saimaa ringed seal	2782	Pup	F	Right	x
Baltic ringed seal	168	Adult a	M	Left		x
	168			Right		x
Baltic ringed seal	218	Unknown	F	Right		x
Baltic ringed seal	458	Pup	M	Right	x
Baltic ringed seal	459	Adult a	F	Right			x
Baltic ringed seal	525	Adult	F	Left		x
	525			Right	x	x
Baltic ringed seal	527	Adult	M	Left		x
	527			Right	x	x
Baltic ringed seal	528	Subadult	M	Right	x	x
Baltic ringed seal	529	Adult	F	Left		x
	529			Right	x	x
Baltic ringed seal	530	Subadult	N/A	Left		x
	530			Right	x	x
Baltic ringed seal	532	Adult	F	Right	x	x
Baltic ringed seal	667	Subadult a	M	Right	x	x
Baltic ringed seal	668	Adult a	F	Left	x	x	x
	668			Right		x
Baltic ringed seal	845	Adult a	F	Left			x
Baltic ringed seal	846	Adult a	F	Right	x	x

^a Age estimation was based on body weight.

2.1. Age and measurements

In cases where age determination was based on the canine tooth by Natural Resources Institute Finland, seals <1 year old were considered pups, seals ≥1 year old but <4 years old were considered subadults, and seals ≥4 years old were considered adults. If these data were not available, seals weighing <30 kg were considered pups, seals ≥30 kg but <42 kg as subadults, and seals ≥42 kg as adults. This classification corresponds well with age estimations based on the canine teeth.

Kidneys from eight Saimaa ringed seals and eight Baltic ringed seals were included in the analysis of kidney weight to body weight ratio. Adults and subadults were analyzed separately. When the weight of only one kidney was known, it was doubled for the analyses. Pups and animals with poor body condition (thickness of the blubber notably less than expected for the age and season) were excluded from this part of the study.

2.2. Silicone casts

Silicone casts were used for visualizing the blood vessels and urinary tract of the kidneys. Three types of two‐compound silicones (3M) were used (ExpressTM 2 Light Body Standard Quick, ExpressTM 2 Regular Body Quick and ExpressTM 2 Light Body Flow Quick). These differed in color, viscosity, and hardening time. The silicone was administered with a gun‐like dispenser (Garant, with a 3M mixing tip) until an increase in resistance was felt. Following administration, the casts were left to set for 10–30 min. Finally, the tissues were dissolved using sodium hypochlorite solution (35%, Sigma‐Aldrich, Switzerland).

Silicone casts were made of the ureter, the renal artery, and, in some seals, the capsular plexus of the kidneys (Table 1). Arterial branching patterns were assessed from 18 kidneys of 14 Saimaa ringed seals and 17 kidneys of 11 Baltic ringed seals. The number of calyces was counted manually from urethral casts of 12 kidneys of 11 Saimaa ringed seals and 10 kidneys of 10 Baltic ringed seals, in which the silicone had properly and evenly advanced to all reniculi. An Olympus SZX9 (Olympus Corporation, Japan) stereomicroscope with a Moticam 1080 camera (Motic Microscopes, China) with 10× magnification was used to build a picture of the arterial silicone cast. The image processing of the silicone casts pictures was enhanced using Photopea software (Ivan Kutskir, www.photopea.com).

2.3. Histology

The kidney tissue samples were fixed in 10% formalin, and histological slides were made of three Saimaa ringed seals and one Baltic ringed seal (Table 2). Reniculi were sectioned parallel or transversely to the axis of the papilla. Sections (5–12 μm) were stained with hematoxylin and eosin, Masson's trichrome, and periodic acid‐Schiff stain. Between 6 and 10 slides were examined per animal. Slides were scanned using a BI Histoscanner (Institute of Biotechnology, University of Helsinki), which uses a 4MP CMOS camera with a speed of 130 fps and a resolution of 0.24 μm/pixel with the combined 20X/0.8 NA objective equivalent to 40× magnification. Scanned samples were examined with the 3DHISTECH Case Viewer program.

TABLE 2.

Specimens used for the histological examination.

Species	Identification	Age group	Sex
Saimaa ringed seal	2742	Pup a	F
Saimaa ringed seal	2755	Pup	M
Saimaa ringed seal	2701	Subadult	F
Baltic ringed seal	459	Adult	F

^a Age estimation was based on body weight.

2.4. Statistical methods

In the analysis of the difference in the number of reniculi between Saimaa and Baltic ringed seals, the normality of both groups was tested with Shapiro–Wilk's test. As there was evidence against normality in the Saimaa ringed seal group (p = 0.0100) non‐parametric tests were used. The difference in the number of reniculi between Saimaa and Baltic ringed seals was analyzed using the Mann–Whitney U test. The previously reported number of reniculi in Arctic ringed seals (Munkacsi & Newstead, 1985) was compared to those of both Saimaa and Baltic ringed seals using one‐sample Wilcoxon signed rank test. The analyses were performed with SPSS Statistics Version 29.0.2.0 (SPSS Inc., Chicago, IL).

2.5. Terminology

The anatomical terminology is mostly in accordance with the International Committee on Veterinary Gross Anatomical Nomenclature (2017) and the International Committee on Veterinary Histological Nomenclature (2017). The term reniculus is used for a lobe of a reniculate kidney. As part of the circulatory system anatomy of pinniped kidneys differs from that of terrestrial mammals, the following terms used by Munkacsi and Newstead (1985) were used in this study.

Ventral slit	As the pinniped kidney does not have renal hilum in the accepted sense, the point of entry to the renal sinus is here termed the ventral slit. In this study, hilum refers to the hilum of a reniculus
Renicular artery	Branch of the arteria renalis, which provides blood to a reniculus. Corresponds to arteria interlobares renis. Renicular artery in Munkacsi & Newstead, 1985
Interrenicular veins	Surround the adjacent sides of reniculi; collect blood from interlobular veins. Interrencular veins in Munkacsi & Newstead, 1985
Interrenicular collecting veins	Situated in junctions of every three reniculi; collect blood from interrenicular veins. Interrencular collecting veins in Munkacsi & Newstead, 1985
Marginal subcapsular veins	Situated in the furrows between reniculi; collect blood from subcapsular veins
Subcapsular venous plexus	Part of the venous plexus situated beneath the renal capsule
Pericapsular venous plexus	Part of the venous plexus situated outside the renal capsule

3. RESULTS

In both species, the kidneys extended from the second to the fifth lumbar vertebrae. The right kidney was located slightly more cranially than the left (Figure 1). The renal arteries and veins and the ureter entered the renal sinus through the ventral slit in between reniculi. The slit was situated on the ventral surface of the kidney, near the medial border and caudo‐cranial midpoint. The shape of the slit varied between individuals, as it was situated in between the segments of the kidney that varied in number and size depending on the renal artery's initial branching. The ratio of kidney weight to body weight was 0.56% in adult Saimaa ringed seals (n = 4) and 0.52% in subadults (n = 4). The corresponding values for the adult Baltic ringed seals (n = 5) were 0.40% and 0.49% for the subadults (n = 3) (Figure 2).

FIGURE 1.

Kidneys of a Saimaa ringed seal (specimen number 2673) in situ.

FIGURE 2.

Kidney weights and body weights of eight Saimaa ringed seals and eight Baltic ringed seals.

For reniculi quantification, the number of calyces was counted from the silicone casts (Figure 3). The number of reniculi was significantly (p = 0.003) less in Saimaa ringed seals (n = 12; mean = 84.3; coefficient of variation = 0.17) than in Baltic ringed seals (n = 10; mean = 100.8; coefficient of variation = 0.09) (Figure 4). The reniculi of both kidneys could be counted for one Saimaa ringed seal individual, and the number differed by one reniculus.

FIGURE 3.

Silicone cast of the ureter of a Saimaa ringed seal (specimen number 2702). Note the caudal and cranial main branches joining near the cranio‐caudal midline. Ventral view, upper border of the picture is toward the cranium.

FIGURE 4.

The number of reniculi in 12 Saimaa ringed seals and 10 Baltic ringed seals, showing the group medians.

3.1. Ureter

Calyces were situated in pairs or singly. In a pair, both calyces either had a ureter stem of a similar length, which joined to form a common ureter stem, or the bases of the calyces were so proximal that only a common ureter stem formed. In single calyces, the ureter stem joined a common ureter stem of a calyx pair (Figure 5). The common ureter stems joined in a tree‐like manner, forming five to seven larger branches, which finally joined to form a caudal or cranial main branch (Figure 3). The caudal and cranial main branches connected near the hilum, approximately in the middle of the cranio‐caudal axis of the kidney, from where the ureter emerged from the kidney through the ventral slit, traveling ventrally to the renal artery. Apart from the division to the caudal and cranial main branch, the ureter did not appear to follow consistent patterns. No renal pelvis was identified.

FIGURE 5.

Close‐up of the ureter cast of a Saimaa ringed seal (specimen number 2702). The common ureter stem of a calyx pair (p) was joined by a ureter stem of a single (s) calyx.

3.2. Arterial circulation

The renal artery for each kidney branched from the aorta (Figure 6). Adrenal arteries frequently branched from renal arteries (not shown). In most cases, the renal artery divided once or twice just before entering the kidney, but in some individuals the first division happened close to the aorta. The arterial division did not follow the urethral branching.

FIGURE 6.

Silicone cast of the renal arteries and aorta of a Saimaa ringed seal (specimen number 2702). Note the differing arterial branching patterns (arrows). Aorta (A), right renal artery (dex), left renal artery (sin). Ventral view, upper border of the picture is toward the cranium.

The kidney was divided into two to four, often unequal‐sized segments, which had mostly or entirely autonomous arterial circulation from a larger branch of the renal artery. The four most common branching patterns (types I–IV) were identified (Figure 7). In two Baltic ringed seals, the branching differed from these recurrent patterns. Variation occurred in the prevalence of branching patterns between the species (Figure 8). The branching pattern could also vary between the kidneys of an individual seal. (Figure 6). After these first divisions, the variation in branching was high.

FIGURE 7.

Branching patterns of the renal artery; upper border of the picture is toward the cranium. Saimaa ringed seals expressed types I–IV, and Baltic ringed seals expressed types I and II. Segment sizes varied.

FIGURE 8.

Incidence of renal artery branching patterns in 18 Saimaa ringed seal kidneys (n = 14) and 17 Baltic ringed seal kidneys (n = 11).

In the renal sinus, renal arteries divided repeatedly, finally giving rise to renicular arteries. Two to three renicular arteries entered a reniculus through the hilum. In most cases, the renicular arteries of a single reniculus originated from a branch or branches of the renal artery in proximity to the hilum, and they frequently divided at or just prior to the hilum. Inside the reniculus, renicular arteries traveled in the corticomedullary junction approximately equidistant from each other. They gave rise to arcuate arteries and occasionally to capsular branches (Figure 9). Arcuate arteries, in turn, gave rise to interlobular arteries (Figure 10). In a few arterial casts the silicone reached the glomeruli but did not advance further (Figure 10).

FIGURE 9.

Capsular arterial branches (highlighted by black background) in a Baltic ringed seal (specimen number 668) silicone cast. Ventro‐latero‐cranial view.

FIGURE 10.

Close‐up of the arterial cast of a Baltic ringed seal (specimen number 168). Arcuate arteries gave rise to interlobular arteries. Arcuate artery (aa), interlobular arteries (ia), glomeruli (*).

3.3. Venous circulation

The venous plexus formed a network surrounding the kidney. The pericapsular part of the plexus had a simple, honeycomb‐like pattern in the pups (Figure 11), whereas it became more sinuous in older animals (Figure 12). Rather than forming true renal veins, the renal plexus drained to its ipsilateral branch of the caudal vena cava at least at three to five sites, where multiple veins of the plexus connected to the caudal vena cava: one each at the cranial and caudal poles of the kidney; and one each near the cranio‐caudal midline ventrally, dorsally, and finally in between them for the veins of the renal sinus. Especially at the cranio‐caudal midline, the dorsal and ventral veins, and the veins from the sinus tended to be merged. In addition, lesser random veins of the plexus were infrequently connected to the vena cava.

FIGURE 11.

Silicone cast of the partly filled veins and arteries of a Saimaa ringed seal (specimen number 2673). The pericapsular part of the venous plexus was well defined, whereas the veins of the renal sinus and renal parenchyma formed a complicated structure. Aorta (A), right branch of the caudal vena cava (dex), left branch of the caudal vena cava (sin), pericapsular venous plexus (*) and veins of the renal parenchyma and sinus (arrowheads). Ventral view, the left border of the picture is toward the cranium.

FIGURE 12.

Silicone cast of the partly filled pericapsular venous plexus of a Saimaa ringed seal (specimen number 2697). Left branch of the caudal vena cava (sin) and pericapsular venous plexus (*). Ventral view; right border of the picture is toward the cranium.

Due to the remarkable diameter and loose, thin‐walled structure of the veins, combined with the great amount of blood in the seal specimens, the venous circulation of the kidney was difficult to study by dissection. Because of the mentioned attributes, venous silicone casts could only be attempted from the kidneys in situ, but still the silicone did not flow well within the venous circulation. Due to the veins' large stretching capacity, the silicone began solidifying before proper progression. Progression in the subcapsular venous plexus in particular was non‐existent, and the silicone rather advanced to the interrenal collecting veins.

3.4. Histology

Histological samples from two Saimaa ringed seal pups, one subadult and one adult Baltic ringed seal were examined (Table 2). All samples were affected by freezing (except 2755) and autolysis. Severely affected samples were omitted from the analyses.

A reniculus corresponded to a unipyramidal kidney or a lobe, consisting of a cortex, medulla, and calyx. The border between adjacent reniculi was marked by interrenicular veins and, in young individuals, occasionally by a thin layer of connective tissue (Figure 12). Apart from the venous system, the structure of a reniculus was mostly typically mammalian.

In the renal sinus, the veins and branches of the ureter, renal artery, and nerve bundles advanced in loose connective tissue (Figure 13). Ureter branches were delineated by a layer of transitional epithelium, surrounded first by a loose layer of connective tissue, followed by a dense layer of connective tissue. In the calyx, the thicknesses of the transitional epithelium and loose connective tissue were reduced. In places where the calyx joined the reniculus, only the dense connective tissue layer overlaid a thin transitional epithelium. The connective tissue layer penetrated to the parenchyma of the reniculus along the corticomedullary border.

FIGURE 13.

Junction of three adjacent reniculi. The border between adjacent reniculi is pronounced with arrowheads. Cortex (c), corticomedullary border (b), medulla (m), interrenicular collecting vein (icv), renicular artery (ra), and interlobar vein (iv). Saimaa ringed seal (specimen number 2755), Masson's trichrome.

The inner lining of arteries appeared folded (Figure 13). They were surrounded by a thick layer of smooth muscle cells and a moderate amount of dense connective tissue. Especially in the larger arteries, the visible amount of collagen intermingled in the layer of smooth muscle cells. Arteries both in the renal sinus and in the reniculi were often accompanied by nerve bundles. The veins were separated from their surroundings only by the endothelium and its basal lamina. The basal lamina appeared thicker in the larger veins.

Interlobular veins had no connective tissue surrounding them. Interrenicular veins collected blood from the interlobular veins and drained to the interrenicular collecting veins situated between every third reniculus (Figure 12). Interrenicular collecting veins, in turn, connected to the subcapsular venous plexus on the kidney surface. Interrenicular veins and interrenicular collecting veins were surrounded by a small amount of connective tissue adjacent to the basal lamina. Interlobular veins directed toward the free face of the reniculus turned toward the nearest edge of the reniculus when they reached its surface. On the edge of the reniculus, they connected to marginal subcapsular veins (Figure 14), which followed the grooves between the reniculi and drained to greater veins of the venous plexus above them.

FIGURE 14.

Renal sinus. A branch of the ureter (u), veins (v), a branch of the renal artery (a), nerve bundles (n). Saimaa ringed seal (specimen number 2755), hematoxylin and eosin staining.

Structures resembling arcuate veins and interlobar veins were observed (Figures 12, 14, and 15). These veins were situated next to some arcuate arteries and renicular arteries, respectively, and they were often embedded in the connective tissue accompanying the arteries. Due to the limited histological sections, it was not possible to confirm where they drained.

FIGURE 15.

Arcuate arteries (aa), renicular arteries (ra), presumed arcuate veins (av), and interlobular veins (iv) surrounded the medulla. Cortex (c), medulla (m), subcapsular veins (arrows), marginal subcapsular vein (msv). Saimaa ringed seal (specimen number 2755), Masson's trichrome staining.

4. DISCUSSION

Both studied species had significantly fewer reniculi (p = 0.002 in Saimaa ringed seals, p = 0.005 in Baltic ringed seals) than previously reported from the Arctic ringed seal (Munkacsi & Newstead, 1985). Furthermore, the Saimaa ringed seal had significantly fewer reniculi than the Baltic ringed seal (Figure 4). The smaller number of reniculi may be argued to be an adaptation to the lower salinity of the Saimaa and the Baltic ringed seal habitats compared to that of marine ringed seals. The coefficient of variation of the number of reniculi was greater in Saimaa ringed seals; however, and the largest number of reniculi (109) in Saimaa ringed seals was close to that (114) recorded from the Baltic ringed seal (Figure 4). The freshwater environment may, therefore, allow more flexibility in the number of reniculi than does brackish water. In the study by Dragert et al. (1975), the coefficient of variation in the number of reniculi in nine harp seals (with one outlier excluded) was 0.1, which is close to our results in Baltic ringed seals. We note that the ambiguous shape of closely situated, poorly filled, or overfilled calyces in some specimens may have caused some error in our analyses. Also, Munkacsi and Newstead (1985) did not specify how many animals the approximate number of reniculi was based on in the Arctic ringed seal. It was possible to count the reniculi of both kidneys in one Saimaa ringed seal, and the number differed by one reniculus. This is in accordance with findings of Dragert et al. (1975) in the harp seal, where the reniculi number between the kidneys of an individual differed by zero to two reniculi.

Although the smaller number of reniculi did not appear to result in lower weight of the kidneys in the Saimaa ringed seals (Figure 2), more data are needed to confirm this finding. In Baltic ringed seals, blood loss due to bullet wounds may have decreased the weight of the kidneys. Concerning the ratio of kidney weight to body weight, the seasonal fluctuation in body weight (Auttila et al., 2016; Kauhala et al., 2018) was not addressed in the analysis, and, in the case of two subadult Baltic ringed seals, the bodyweight was an estimate provided by the hunters to Natural Resources Institute Finland. Furthermore, despite excluding animals with poor body condition, the naturally deceased Saimaa ringed seals presumably had an ongoing disease process, which could have affected their bodyweight. Nonetheless, when the absolute weights of the kidneys were compared between the species, three out of four Saimaa ringed seals' kidneys of both adult and subadult animals weighed more than the Baltic ringed seals' in respective age groups (Figure 1). Further studies are warranted to investigate whether the microstructure of a reniculus differs between species, for example, in the number or size of the glomeruli or in the length of the tubulae. Considering other vertebrates, some fish species with populations living in different osmotic environments show changes in glomerular size and kidney gene expression (Hasan et al., 2017).

In both species there were no apparent renal pelvis, as the diameter of the ureter was greatest when emerging from the ventral slit. This finding is in accordance with previous studies in other pinnipeds (Guzsal, 1959, Bester, 1975, Dragert et al., 1975, Vardy & Bryden, 1981).

Slijper (1958) and Williams and Worthy (2002) remarked that the number of reniculi correlates roughly with the capacity for urine excretion. The physiology of freshwater seal kidneys was assessed by Hong et al. (1982), who compared the freshwater Baikal seal (Pusa sibrica Gmelin, 1788) to the Arctic ringed seal. They concluded that Baikal seal renal functions (e.g., urine concentration ability) did not markedly differ from Arctic ringed seal kidney function (Hong et al., 1982). Baikal seals did, however, conserve sodium better than Arctic ringed seals did during a fasting experiment (Hong et al., 1982). This finding correlated with the low sodium availability in the Baikal seal's habitat (Hong et al., 1982).

There is little data on reniculi numbers in pinnipeds (Dragert et al., 1975; Munkacsi & Newstead, 1985; Vardy & Bryden, 1981). This may partly be due to the lack of connective tissue between the reniculi, which renders counting by dissection laborious. In our study, using silicone to visualize the kidney structures made counting the reniculi easier, as the urethral model could be manipulated quite freely without breaking it. However, as the silicone's progression within the kidney could not be properly evaluated during administration, for multiple samples we had stopped before the silicone had reached all the calyces. Therefore, some casts could not be used in counting the calyces. On the other hand, overfilling caused ruptures in the tissues.

Regarding the circulatory system, our findings were mostly in accordance with earlier descriptions. However, we did find veins resembling arcuate and interlobar veins (Figures 15 and 16), which implied that these animals possess both extrinsic and intrinsic venous systems. Due to the thin‐walled and loose structure, these veins were challenging to document.

FIGURE 16.

Corticomedullary border. Cortex (c), medulla (m), arcuate artery (aa), interlobular artery (ia), presumed arcuate vein (av), nerve bundle (n). Saimaa ringed seal (specimen number 2755), Masson's trichrome.

Between the Saimaa and Baltic ringed seals, the circulatory anatomy of the kidneys did not differ remarkably. The renal artery's branching pattern type II corresponded to Guzsal's (1959) description in the one studied specimen of the Ladoga ringed seal. Partially differing branching patterns of the renal artery between the Saimaa and Baltic ringed seals (Figure 7) are unlikely to affect kidney physiology. Similar minor differences between the species have been reported from their cranial circulation (Laakkonen et al., 2024). Together with the varying anatomy of the venous circulation, our results more likely emphasize the flexibility of kidney circulation development (Figure 7).

The primary limitation of our study was the small sample size. Because the Saimaa ringed seal is an endangered species, data collection was difficult, especially from the adult animals. Most Saimaa ringed seals in this study went through a pathological examination, after which the sample material was divided between multiple research groups. Therefore, it was not always possible to, for example, acquire a whole kidney for the purposes of this study. The stage of autolysis was affected by the time between death and freezing, and the season and weather at the time of death, along with the cause of death. The best‐preserved Saimaa ringed seals were typically pups and subadults, which die in fishing nets more commonly than adults. As all Saimaa ringed seals (apart from specimen number 2755) and Baltic ringed seals were frozen and thawed before sample taking, storage conditions may have affected the histological quality of our material.

AUTHOR CONTRIBUTIONS

Heini Nihtilä: Conceptualization; investigation; writing – original draft; methodology; visualization; writing – review and editing; validation; software; formal analysis; data curation. Juha Laakkonen: Investigation; conceptualization; writing – review and editing; methodology; validation; project administration; resources; supervision.