Physiological Solutions

PHYSIOLOGICAL SOLUTIONS, solutions in which a living cell finds physiological, i.e., conditions closest to normal; the main conditions are: 1) osmotic pressure; 2) the quantitative ratio of the most important ions (H, OH, K, Ca, Na, Mg; in some newer physiological solutions also Cl, PO₄, CO₃ and SO₄); 3) the correct ratio of dissolved gases (O₂ and CO₂); 4) the reduction potential; 5) the colloidal-chemical state (the presence of colloids in an amount ensuring the emergence of a membrane potential normal for a given cell or tissue) and 6) buffer properties (connected with the previous factors). A specific pH of physiological solutions is also connected with the ratio of ions and buffer substances. Pfeffer and Nägeli first showed that cells perish in distilled water as a result of plasmolysis (hemolysis); at the same time, in surgical practice, it turned out that in cases of great blood loss, the infusion of a 0.6–1% solution of sodium chloride often acted as a lifesaver, while the infusion of distilled water led to death. The first physiological solutions were only isotonic, i.e., they ensured only normal osmotic pressure (from 5 to 8.5 atmospheres). Physiological solutions found their main medical application in the form of subcutaneous, intravenous, and other injections; they are widely used in the technique of research work with surviving isolated organs (see Isolated Organs, Tissue Cultures). The credit for introducing saline solutions for work with isolated organs belongs to Ringer (1882). Ringer, in his classic experiments, showed that the simultaneous addition to an isotonic solution of table salt of a small amount of calcium chloride and potassium chloride maintains the work of an isolated frog heart for a long time. This laid the foundation for so-called "balanced" solutions, which are part of artificial physiological solutions. To Ringer also belongs the introduction into the composition of the solution he proposed (see Ringer's solution) of sodium bicarbonate, which gives the solution the necessary alkalinity. Tyrode showed that for uniform contractions of isolated smooth-muscle organs, it is useful to add magnesium chloride to the nutrient fluid. The composition of Tyrode's solution also includes monobasic sodium phosphate (NaH₂PO₄), which, along with NaHCO₃, ensures the buffer properties of physiological solutions: Aq. destillatae 1000.0; NaCl 8.0; KCl 0.2; CaCl₂ 0.2; MgCl₂ 0.1; NaHCO₃ 0.1; NaH₂PO₄ 0.05; some laboratories add 1.0 of grape sugar to this. Cushny proposes the same recipe for Tyrode's fluid but uses crystalline CaCl₂. In the laboratory of Magnus (Utrecht), the following order of preparing Tyrode's fluid is adopted: 80.0 NaCl; 20 cm³ of a 10% KCl solution; 20 cm³ of a 10% CaCl₂ solution; 10 cm³ of a 10% MgCl₂ solution are diluted in 9 liters of water; 1 liter of water, in which 10.0 g of NaHCO₃ and 10 cm³ of 5% NaH₂PO₄ are dissolved, is slowly added to them while stirring. Ringer's solutions, like other similar solutions, must be prepared from chemically pure reagents in water distilled from glass or porcelain vessels to avoid the toxic effect of traces of heavy metals, which are often present in ordinary water distilled from metal stills. To avoid the precipitation of calcium in the form of carbonate, it is recommended to dissolve calcium chloride and soda separately, mixing only their cooled, diluted solutions. When using crystalline calcium chloride instead of anhydrous, the amount of water of crystallization should be taken into account (i.e., take 1.97 g of CaCl₂·6H₂O instead of 1 g of CaCl₂ siccum). Various authors give different recipes for saline nutrient fluids. We present some of them. Variants of Ringer's solution for frog organs: 1) Clark—NaCl 0.7%; KCl 0.014%; CaCl₂ (siccum) 0.012%; NaHCO₃ 0.02%. Bayliss gives the same recipe, but with 0.65% NaCl; 2) Cushny—NaCl 0.6%; KCl 0.0075%; CaCl₂ (cryst.) 0.026%; NaHCO₃ 0.01%. Ringer's solutions for warm-blooded animals: 1) Rush (Langendorff lab)—NaCl 0.8%; KCl 0.0075%; CaCl₂ 0.01%; NaHCO₃ 0.01%; 2) Trendelenburg—NaCl 0.8%; KCl 0.0075%; CaCl₂·6H₂O (cryst.) 0.02%; NaHCO₃ 0.01%. Variants of Locke's solutions (for warm-blooded animals): 1) Locke (second variant)—NaCl 0.9%; KCl 0.02%; CaCl₂ (siccum) 0.02%; NaHCO₃ 0.02%; Sacch. uvic. 0.1%. This variant of Locke's solution was used in Kravkov's laboratory during his numerous works with isolated organs. 2) Cushny (first variant)—NaCl 0.92%; KCl 0.042%; CaCl₂·6H₂O (cryst.) 0.024%; NaHCO₃ 0.015%; Sacch. uvic. 0.1%. 3) Dale—NaCl 0.9%; KCl 0.042%; CaCl₂ (siccum) 0.024%; NaHCO₃ 0.05%; MgCl₂ 0.0005%; Sacch. uvic. 0.05%. (Cushny gives the same recipe—second variant—but indicates crystalline CaCl₂·6H₂O instead of anhydrous). Variants of Tyrode's solution: 1) Sollmann—NaCl 0.8%; KCl 0.02%; CaCl₂ (siccum) 0.01%; MgCl₂ 0.01%; NaHCO₃ 0.1%; NaH₂PO₄ 0.005%; Sacch. uvic. 0.1%; 2) Trendelenburg—NaCl 0.8%; CaCl₂·6H₂O (cryst.) 0.04%; KCl 0.02%; MgCl₂ (cryst.) 0.02%; NaHCO₃ 0.1%; NaH₂PO₄·H₂O 0.005%. He recommends the following order of preparation. Prepare separately the "first solution" (Tyrode I): dissolve 100.0 NaCl in 10 liters of water and add 50 cm³ of a 10% solution of crystalline CaCl₂·6H₂O, 20 cm³ of a 10% KCl solution, and 20 cm³ of a 10% MgCl₂ solution; "second solution" (Tyrode II): dissolve 10.0 NaHCO₃ in 2 liters of water and, following dissolution, add 0.5 NaH₂PO₄ + H₂O. Tyrode II must be kept in bottles filled to the top. For the experiment, 200 cm³ of the second solution is added to 800 cm³ of the first solution. The finished solution should not remain open for more than two hours, otherwise the release of CO₂ is facilitated, the reaction becomes alkaline, and calcium carbonate may precipitate. Trendelenburg reports that instead of distilled water in experiments with segments of the uterine horn, he used tap water, which contains very little calcium in Freiburg. The laboratory of S. V. Anichkov (Leningrad) has the same experience regarding Neva tap water, which, after being filtered through a Chamberland filter, was successfully used for Locke's fluid when working with isolated ears and hearts; however, in some periods, the tap water proved to be clearly unsuitable.

Numerous studies have been devoted to the question of the significance of various components and properties of "balanced" solutions for the survival of organs. The most essential conditions are the following: 1) the osmotic pressure of the nutrient fluid (mainly the concentration of NaCl) must be isotonic to the tissues of the given animal species. Thus, for the frog, it is lower (0.6% NaCl), for warm-blooded animals higher (from 0.8% to 0.9% NaCl). True, some deviation from isotonia is permissible, in particular a slight excess of it. In Sollmann's opinion, with hypotonia of the nutrient fluid, the function of isolated organs increases, but the duration of their survival is shortened; hypertonic solutions have the opposite effect. 2) The balance of potassium and calcium ions is one of the basic conditions for maintaining the vital activity of tissues. Experience has shown that some variations in the amount and ratio of potassium chloride and calcium chloride included in the nutrient fluid are permissible. Locke himself, for example, pointed out that an isolated rabbit heart works just as well with the second variant of his fluid (CaCl₂ 0.02%; KCl 0.02%) as with the first variant (CaCl₂ 0.024%; KCl 0.042%). In various formulas of Ringer's solutions, the content of calcium chloride fluctuates from 0.01% to 0.026%, and potassium chloride from 0.0075% to 0.05%. Research shows that a change in the ratio of potassium and calcium undoubtedly affects the functional characteristics of organs (sensitivity to poisons, electrical excitability, etc.), and therefore, in experiments of the same series, a fluid with a strictly constant content of potassium and calcium must be used. 3) The reaction of the fluid (pH) must be close to that of blood, i.e., slightly alkaline (pH = about 7.4); at the same time, the nutrient fluid, like blood, must possess buffer properties and neutralize the acids formed in the tissues.

In NaHCO₃ solutions, the buffer system is the bicarbonate system, and their ratio determines the pH value of the fluid. At a given concentration of NaHCO₃, the pH of Ringer's solution depends on the amount of carbonic acid dissolved in the water in which the solution is prepared. Phosphates, which provide very satisfactory pH stability, are part of Tyrode's fluid (see above). Significant deviations in pH, both in the acidic and alkaline directions, have a sharp effect on the function of isolated organs. Clark showed that when the pH of Ringer's solution is changed from 7.6 to 6.5, the amplitude of frog heart contractions drops several times. According to the research of N. S. Spassky, the optimum pH for a frog's heart lies between 6.9 and 7.6, and for a rabbit's heart between 7.1 and 7.45; cardiac arrest in the frog occurs at pH=6.0 and above 8.25; for the rabbit, these limits are 6.3 and 7.7–7.85. It should be borne in mind that even with the same composition of the fluid, its reaction can change under the influence of various factors that cause the release of CO₂ (standing of an open solution, shaking with air, heating, passing oxygen, etc.), which is why pH should be monitored in precise experiments. 4) To bring the living conditions of the tissues of an isolated organ closer to normal, maximum saturation of the nutrient fluid with oxygen is necessary, which is especially essential when working with isolated organs of warm-blooded animals. This saturation is usually carried out by constantly passing a stream of oxygen through the nutrient fluid throughout the entire experiment. Some laboratories, when working with smooth muscle organs, use a stream of air instead of pure oxygen. It is necessary to keep in mind that when oxygen is passed through a column of physiological solution, along with O₂ saturation, CO₂ is extracted from it, due to which the pH of the fluid gradually increases. To maintain the stability of the fluid's pH, it is most rational to pass not pure oxygen, but a mixture with CO₂. In Magnus's laboratory, it is customary to pass the stream of air intended for the fluid through a NaHCO₃ solution beforehand. Constant passing of oxygen can be replaced by preliminary saturation of the fluid with it before the experiment by passing a stream of O₂ through it for a sufficiently long time. Such saturation, in order to avoid significant disturbance of the pH, is better performed before adding NaHCO₃ to the fluid. Stronger oxygen saturation can be obtained when the fluid is in an atmosphere of oxygen under increased pressure. This is achieved in the apparatus of the Rostov Physiological Institute, in which the pressure of the oxygen located in the flask with the nutrient fluid serves as the force ensuring the perfusion of the fluid through the organ. But even under these optimal conditions, the oxygen content in Ringer-Locke fluid is significantly less than in arterial blood, and thus the organs irrigated by it are in conditions of relative oxygen starvation. The most careful observance of these main conditions does not, however, give 'balanced' solutions complete physiological identity with blood. There have been proposals to add gum arabic (Bayliss) or gelatin (N. N. Savitsky) to Ringer's solution to give it the colloidality corresponding to blood. But these preparations usually contain impurities that are not indifferent to tissues, and the proposed method has not gained widespread use. Consequently, the saline solutions used for isolated organs do not create a completely physiological environment for tissues. When using formulas containing Ca and Mg salts along with NaHCO₃ or phosphates, the physiological solution is prepared first without carbonates and phosphates, sterilized, and then, after cooling, the NaHCO₃ and phosphate solution is added. In medical institutions with modern equipment, sterilization of the finished physiological solution should be carried out in autoclaves filled with CO₂ under a pressure of two atmospheres (from a cylinder). If it is necessary to prepare a complex physiological solution with the addition of medicinal substances, the composition must be calculated taking into account the osmotic pressure created by the medicinal substance itself. It is most convenient to perform this calculation using the Oberhard formula: an isotonic solution is obtained by dissolving 1/1 k · M grams of the substance in 1 liter, where M means the molecular weight of the substance, k is a conventional coefficient, equal to unity for strong electrolytes (NaCl, KCl), 1.4 for weak electrolytes (NaHCO₃, Na₂HPO₄, most alkaloid salts), and 2.0 for non-electrolytes (glucose, sugar, urea, urotropin, etc.). According to this formula, for example, a solution of 8.35 g of NaCl in 1 liter (the Pharmacopoeia VII gives the figure 8.5 g in 1 liter) or a solution of 51.4 g of glucose (M=180) in 1 liter (rounded off = 5%), etc., will be isotonic. A complex solution is considered as a mixture of individual physiological solutions; for example, wishing to prepare a 1% urotropin solution (M=140), one calculates that a solution of 40 g of urotropin in 1 liter (i.e., about 4%) will be isotonic, and therefore it must be diluted fourfold (to obtain 1% urotropin) with an isotonic solution of table salt; in the end, one must prescribe: Urotropini 40.0; Natr. chlorati (calculated for 3 liters) 25.5; Aq. dest. ad 4000 cm³ or proportionally less, e.g., Urotropini 1.0; Natr. chlorati 0.64; Aq. dest. ad 100 cm³. S. Apichvoye. For therapeutic purposes, a simple physiological solution of NaCl is most often used, which is a solution of 7.5–9.0 NaCl per 1000.0 of distilled water. It is used: 1) to compensate for the body's loss of water (profuse bleeding, dehydration in the algid stages of cholera and in cholera-like diarrhea); 2) to increase dropped blood pressure (collapse during anesthesia, surgical shock), often with the addition of adrenaline; 3) to improve blood circulation in acute infectious diseases, in peritonitis; 4) for washing the body in infectious (typhoid, sepsis, etc.) and other toxemias, as well as in various toxicodermias and dermatoses (general skin itching, urticaria, some forms of chronic eczema, polymorphic exudative erythema, etc.); the effect of the saline solution is noticeably increased if its administration is combined with preliminary bloodletting of 100–200 cm³ of blood; 5) to quench thirst when this cannot be done in the usual way (after operations, in diseases of the oral cavity, pharynx, esophagus, with persistent vomiting, etc.). Physiological solution is most often administered by subcutaneous or intravenous infusions. Usually, 500–1000 cm³ of solution with a temperature of 38–39° are infused; the simultaneous administration of large quantities (1.5–2 liters) can be accompanied by the appearance of oliguria, edema, and asystole (due to overloading the body with water and salt). Subcutaneous infusions are more painful than intravenous ones, but their effect is more prolonged. When using infusion, it is necessary to carefully sterilize both the equipment and the solution. Often, physiological solution is administered per rectum in the form of drip enemas, and by this method, up to 5 liters can be administered per day; the effect with it is even slower and more gradual, and there is no danger of overloading the body. This method is easily applicable, does not require special sterilization, is painless, and is tolerated by the patient for several hours without unpleasant sensations. Sometimes physiological solution is introduced into the abdominal cavity—during laparotomy, either directly or through a drain left in the wound, and in pediatric practice by puncture of the abdomen. In some cases, physiological solution is used locally, mainly for neuralgias, and it is injected in small quantities (10–100 cm³) into the nerve itself. When using a physiological solution of NaCl, mainly during its infusion, a subsequent (salt) fever is sometimes observed, which may partly depend on the use of not freshly distilled water. The addition of Ca salts to the NaCl solution prevents the appearance of salt fever, which is why sometimes, instead of a physiological solution of NaCl, the above-mentioned more perfect physiological solutions are used, and especially normosal (see), which is very convenient for the rapid preparation of a solution (the contents of 1 ampoule are dissolved in 1 liter of freshly distilled water). A contraindication to the use of physiological solution can only be a sharp disorder of cardiac activity with phenomena of congestion, cyanosis, and pulmonary edema. Recently, in order to increase the body's nutrition and reduce metabolic toxicosis (azotemia, ketonemia), an isotonic (5%) glucose solution is very often used; it is administered by intravenous, subcutaneous, or rectal infusions (100–500 cm³) in a form warmed to body temperature.

A. Molchanov. The term 'Physiological solutions' is sometimes applied to nutrient fluids serving to maintain the vital functions, or 'nourishment,' of surviving isolated organs (see Isolated organs). The simplest and at the same time widely used fluids serving for the irrigation of isolated organs are, however, not nutrient in the direct sense of the word, since in the majority of cases they do not carry nutrient material for the tissues. Nutrient fluids can be divided into two groups: 1) blood preparations, 2) artificial fluids based on saline solutions. The blood of the same species of animal from which the isolated organ was taken would, of course, be the most perfect nutrient fluid for it, but the coagulability of whole blood makes it impossible to use it as such outside the organism. Hence the numerous attempts to use non-coagulating blood preparations for the nourishment of isolated organs. Even the first authors who conducted systematic experiments on isolated organs widely used defibrinated blood; however, it has significant drawbacks, since the products contained in it, which are formed during coagulation, are far from indifferent to the tissues; in particular, they cause vascular spasm and spastic contraction, and subsequently paralysis of smooth muscles. Starling showed that the toxic properties of defibrinated blood are significantly reduced by its constant circulation through the vessels of an isolated lung; consequently, perfusion of isolated organs included in Starling's 'heart-lung' preparation with defibrinated blood is one of the best methods for maintaining their vital functions. Many times, attempts have been made to add substances that prevent coagulation to the blood used for organ perfusion. Salts that prevent blood coagulation (nitrates, oxalates, etc.) are poorly suited for this purpose, as they sharply disrupt the ionic balance. Substances such as hirudin and heparin are more suitable. But even hirudinized and heparinized blood are not ideal nutrient fluids, proving in most cases not entirely indifferent to the tissues. Apparently, the addition of these substances, while delaying blood coagulation, does not entirely prevent the formation of certain toxic substances in the blood. Both defibrinated blood and non-coagulating blood preparations are not ideal nutrient fluids; however, they are still superior to saline solutions, providing the organs with conditions closer to natural ones. This is especially evident in experiments on the organs of warm-blooded animals with the investigation of the tissue metabolism of isolated organs. Saline solutions used as nutrient fluids have a great advantage in the simplicity, definiteness, and constancy of their composition.